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In rodents, brown adipose tissue (BAT) regulates cold- (CIT) and diet-induced thermogenesis (DIT). Whether BAT recruitment is reversible and how it impacts on energy metabolism has not been investigated in humans. We examined the effects of temperature acclimation on BAT, energy balance and substrate metabolism in a prospective crossover study of 4-month duration, consisting of 4 consecutive blocks of 1-month overnight temperature acclimation [24°C (month 1) → 19°C (month 2) → 24°C (month 3) → 27°C (month 4)] of five healthy men in a temperature-controlled research facility. Sequential monthly acclimation modulated BAT reversibly, boosting and suppressing its abundance and activity in mild cold and warm conditions (p<0.05), respectively, independent of seasonal fluctuations (p<0.01). BAT-acclimation did not alter CIT but was accompanied by DIT (p<0.05) and post-prandial insulin sensitivity enhancement (p<0.05), evident only after cold-acclimation. Circulating and adipose tissue, but not skeletal muscle, expression levels of leptin and adiponectin displayed reciprocal changes concordant with cold-acclimated insulin sensitization. These results suggest regulatory links between BAT thermal plasticity and glucose metabolism in humans, opening avenues to harnessing BAT for metabolic benefits.
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1
Temperature-acclimated brown adipose tissue modulates insulin sensitivity in 1
humans 2
3
Paul Lee
1
, Sheila Smith
1
, Joyce Linderman
1
, Amber B Courville
2
, Robert J Brychta
1
, 4
William Dieckmann
3
, Charlotte D Werner
1
, Kong Y Chen
1
, Francesco S Celi
1,4
5
6
Diabetes, Endocrinology, Obesity Branch, National Institute of Diabetes and 7
Digestive and Kidney Diseases
1
, Department of Nutrition, Clinical Center
2
, PET 8
Department, Clinical Center
3
, National Institutes of Health, Bethesda, USA and 9
Division of Endocrinology and Metabolism
4
, Virginia Commonwealth University, 10
Richmond, USA 11
12
Correspondence author: Francesco S Celi, Division of Endocrinology and 13
Metabolism
4
, Virginia Commonwealth University, 1101 East Marshall Street, Sanger 14
Hall, Room 7-007, Richmond, USA 15
Tel: 1 804 828 Email: fsceli@vcu.edu 16
17
None of the authors have any conflicts of interest to disclose 18
19
Abbreviated title: Temperature-acclimated human brown adipose tissue 20
Key words: white adipose tissue, beige adipose tissue, thermogenesis, adiponectin, 21
leptin, CIDEA, GLUT4 22
23
Word count: 177 (Abstract); 3999 (Main Text) 24
Number of tables: 4 Number of figures: 4 Supplemental Data: 1 25
26
Page 1 of 59 Diabetes
Diabetes Publish Ahead of Print, published online June 22, 2014
2
Abstract 27
28
In rodents, brown adipose tissue (BAT) regulates cold- (CIT) and diet-induced 29
thermogenesis (DIT). Whether BAT recruitment is reversible and how it impacts on 30
energy metabolism has not been investigated in humans. We examined the effects of 31
temperature acclimation on BAT, energy balance and substrate metabolism in a 32
prospective crossover study of 4-month duration, consisting of 4 consecutive blocks 33
of 1-month overnight temperature acclimation [24°C (month 1) 19°C (month 2) 34
24°C (month 3) 27°C (month 4)] of five healthy men in a temperature-controlled 35
research facility. Sequential monthly acclimation modulated BAT reversibly, boosting 36
and suppressing its abundance and activity in mild cold and warm conditions 37
(p<0.05), respectively, independent of seasonal fluctuations (p<0.01). BAT-38
acclimation did not alter CIT but was accompanied by DIT (p<0.05) and post-prandial 39
insulin sensitivity enhancement (p<0.05), evident only after cold-acclimation. 40
Circulating and adipose tissue, but not skeletal muscle, expression levels of leptin and 41
adiponectin displayed reciprocal changes concordant with cold-acclimated insulin 42
sensitization. These results suggest regulatory links between BAT thermal plasticity 43
and glucose metabolism in humans, opening avenues to harnessing BAT for 44
metabolic benefits. 45
46
47
48
49
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Introduction 50
51
Unhealthy diet and physical inactivity are the major culprits to the obesity crisis, 52
although other environmental factors may also contribute (1). An overlooked 53
component in energy balance is adaptive thermogenesis, which comprises diet-54
induced thermogenesis (DIT) and cold-induced thermogenesis (CIT). DIT is the 55
portion of energy expended following food ingestion, beyond the energy cost of 56
digestion/absorption (2). The CIT response defends core temperature during cold 57
exposure (3). In rodents, both processes are chiefly regulated by brown adipose tissue 58
(BAT). Through the action of uncoupling protein 1 (UCP1), energy is converted into 59
heat, and represents a form of energy expenditure (EE) as energy is dissipated to the 60
environment. BAT stimulation protects animals against diet-induced obesity and 61
glucose intolerance (4). 62
63
In addition to “classic BAT” in the interscapuar region, cold exposure also induces the 64
emergence of brown adipocyte-like cells (beige/brite adipocytes) within white adipose 65
tissue (WAT) in animals (5; 6). Brown/beige fat generates heat from glucose/lipids 66
and their high substrate utilization underlies protection against diet-induced insulin 67
resistance in genetic, pharmacological and/or transplantation models of invigorated 68
brown/beige fat status (7-9). In humans, histological examination had demonstrated 69
the presence of BAT in adult in the 1970-80’s (10-12), although BAT whole body 70
abundance was not fully appreciated until its visualization was made possible by 71
Positron Emission Tomography (PET)/CT (13-17). Not only is BAT inducible in 72
humans (18; 19), it also exhibits oxidative capacity (20) and classic BAT/beige fat 73
features (21; 22), thus forming the basis for the quest of BAT/beige fat-enhancing 74
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4
strategies as anti-obesity treatments (23). 75
76
Acute cold exposure (hours) increases BAT activity (13; 15-17; 24), while longer-77
term exposure (days/weeks) expands BAT volume (25; 26). Because BAT 78
recruitment could reduce adiposity (26), it suggests BAT may impact whole body 79
energy homeostasis. The corollary is that reduced cold exposure could suppress 80
BAT/beige fat function in humans, with potential obesogenic consequences (27). To 81
date, cold exposure is the best-known activator (15-17) and recruiter (25; 26) of BAT, 82
and associative data have linked higher BAT abundance with leanness and lower 83
glycemia in humans (13; 15; 28; 29). Whether BAT withers under warm exposure and 84
if BAT recruitment triggers compensatory metabolic and/or behavioral adaptations 85
have not been investigated, but are integral to BAT physiology. Rodent studies have 86
revealed a complex interplay between housing temperature, BAT recruitment and 87
energy balance, which ultimately determines metabolic phenotype (30). To better 88
appreciate the metabolic significance of human BAT, and the implications of BAT 89
status on health, BAT recruitment interventions should be examined in the context of 90
whole body energy metabolism. 91
92
In this study, we investigated the effects of long-term mild cold and warm exposure 93
by minimal overnight manipulation of ambient temperature on individual BAT status 94
and the corresponding energy/substrate homeostatic responses. We hypothesize 95
human BAT exhibits plasticity and its activity modulates systemic energy 96
metabolism. 97
98
99
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METHODS 100
101
Subjects 102
Five healthy men were recruited through local advertisement and provided written 103
informed consent. NIDDK-NIAMS IRB approved the study (ClinicalTrials.gov: 104
NCT01730105). Supplemental Figure S1 summarizes recruitment, allocation and 105
intervention. 106
107
Overall design 108
This is a prospective crossover study consisting of 4 consecutive blocks of 1-month 109
duration [Supplemental Figure S2]: it incorporates i) sequential monthly thermal 110
acclimation over 4 months, and ii) acute thermo-metabolic evaluations at the end of 111
each study temperature regime. Volunteers were admitted to the Clinical Research 112
Unit (NIDDK) in Bethesda, Maryland (April-November 2013) for the entire 4 113
months. 114
115
Monthly thermal acclimation 116
Volunteers resided in a temperature-adjusted private room, engaged in usual daily 117
activities and returned to their room each evening. Room temperature was adjusted in 118
the following sequence: 24°C(month 1)19°C(month 2)24°C(month 119
3)27°C(month 4). Volunteers were exposed to the study temperature for at least 10 120
hours each night, wearing standardized hospital clothing with a combined thermal 121
insulation value of 0.4 (clo). Only bed sheets were provided. Volunteers were asked to 122
not deviate daily activity level over the study period. Each subject therefore acted as 123
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his own control. At 08:00 at the end of each month, volunteers were admitted to a 124
whole-room indirect calorimeter for thermo-metabolic evaluation. 125
126
Temperature monitoring 127
Volunteers wore two temperature data loggers (Extech RHT20, Nashua, NH), one 128
“external to clothing” to track environmental temperature; the other “within clothing” 129
to track immediate temperature changes in the “microenvironment” within clothing. 130
We averaged individual exposed temperature every 30 minutes for the entire 4-month 131
period, allowing us to record environmental temperature variations and “true 132
temperatures” the individual was being exposed to. 133
134
Diet 135
All meals, including pre-packed lunches/snacks, were provided with the following 136
composition: 50% carbohydrate, 20% protein, 30% fat. The first month was an 137
equilibration period, during which volunteers followed a weight maintenance diet. 138
After month 1, subjects ate according to hunger. Caloric/macronutrient content was 139
calculated based on weight maintenance requirements, determined during 140
equilibration month. Any unconsumed foods were returned/weighed for 141
energy/macronutrient intake calculation. Subjects met study dieticians twice weekly 142
to verify food diaries/compliance. Total intake/macronutrient was computed/analyzed 143
using three-dimensional food models (ProNutra, version 3.4.0.0., Viocare 144
Technologies, Princeton, NJ). 145
146
Appetite/hunger assessment 147
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Subjects completed questionnaires assessing appetite twice a week before/after 148
breakfast, by marking on a visual analogue scale (VAS) (10cm long) responses to the 149
questions: 1) How hungry are you? 2) How full are you? 3) How much food can you 150
consume? These questions gauged hunger, satiety and desire to eat before/after meals. 151
152
Before each monthly thermal-metabolic evaluation, volunteers underwent an ad 153
libitum meal test, consisting of a selection of food items displayed in a vending 154
machine. Subjects ate until they felt ‘comfortably full’. Total energy/macronutrient 155
intake were recorded, together with ratings of appetite (hunger, satiety, desire to eat) 156
using the same weekly questionnaire at T=−10, 0, 60, 120, 180, 240 and 300 min 157
where initiation of the meal was defined as T=0 min. 158
159
Acute thermo-metabolic evaluations 160
Thermo-metabolic evaluation was scheduled at the end of each month [Supplemental 161
Figure S2], modeled on our previous published methods (24; 31), with total energy 162
expenditure (EE) calculated as previously described (24; 31). Volunteers underwent 163
two 24-hour sessions in a whole room calorimeter, exposed to first 24°C (day 1) then 164
19°C (day 3), with a resting 24-hour period in between. The temperature order was 165
not randomized because our previous study did not reveal a sequence effect (31). 166
Testing at the two temperatures allowed us to evaluate how monthly acclimation 167
modulated EE/metabolism at both thermoneutral and mild cold conditions. Lunch 168
(Boost Plus, Nestle Healthcare Nutrition, Inc., Vevey, Switzerland) and dinner 169
(selected from Metabolic Menu) were provided at 13:00 and 19:00, consisting of 1/3 170
and 2/3 of daily caloric intake, respectively, based on calculation from equilibration 171
month. CIT was calculated as difference in total EE between 24°C and 19°C, and DIT 172
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as difference in pre- (08:00-13:00) and post-lunch (13:00-19:00) EE. As the test meal 173
carried identical caloric and macronutrient content, we attributed any changes 174
observed to arise from adaptive thermogenesis, because the facultative component 175
(i.e. digestion/absorption) should be relatively unaltered. Shivering response was 176
quantified by surface electromyography (EMG), as previously described (32), and 177
volunteers reported perception to cold each month during EE testing using VAS. 178
Hormonal/metabolic parameters were measured in venous samples. Post-prandial 179
insulin sensitivity was calculated after a mixed meal (33), and adipose resistance 180
index by product of free fatty acid × insulin. At the conclusion of thermo-metabolic 181
study, body composition was measured, as previously published (24; 31). 182
183
PET/CT scanning 184
Positron-Emission Tomography (PET)-Computed Tomography (CT) was performed 185
using Siemens Biograph mCT (Siemens Healthcare, Ill., USA) (32). PET/CT was 186
undertaken at 08:00 the morning after the 19°C testing day, at the end of each 187
acclimation month. Attenuation corrected PET-CT images were analyzed using 188
custom software built with IDL (Excelis Visual Information Solutions, Inc., Boulder, 189
CO). A 3-dimentional region of interest (ROI) was defined cranially by a horizontal 190
line parallel to the base of the C4 vertebra, and caudally by an oblique line traversing 191
the manubriosternal joint and T8 transverse process [Supplemental Figure S3]. BAT 192
was defined as tissue with Hounsfield units -300 to -10 on CT (i.e. fat density) with a 193
lean body mass standardized uptake value (SUV) of 2 (i.e. high glucose uptake). The 194
chosen ROI captures major BAT depots in the cervical, supraclavicular, axillary, 195
superior mediastinal and paravertebral areas. This ROI was chosen because spurious 196
myocardial/renal excretory FDG uptake could not be reliably excluded from BAT. 197
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This approach allowed examination of BAT evolution within a well-defined region of 198
adipose tissue across 4 months. 199
200
PET-CT parameters 201
The following parameters were analyzed: BAT volume, mean SUV and activity. BAT 202
volume, defined as the sum of the volume of all voxels that met HU-SUV criteria, 203
represents activated BAT. Mean SUV (normalized by FDG dose and lean body mass) 204
of ROI represents mean metabolic activity within BAT-harboring region. BAT 205
activity represents the total radioactivity (in MBq) within ROI and captures both 206
changes in volume and mean FDG uptake. Furthermore, because fat exhibits 207
metabolic activity as a continuum and the chosen SUV threshold of 2 is arbitrary, 208
and may potentially exclude more diffuse enhancement of adipose metabolic activity, 209
we also quantified mean SUV in the entire ROI within tissue of fat density (HU: -300 210
to -10). Mean SUV of whole fat depot estimates overall metabolic activity, and may 211
capture both BAT and diffuse beige fat activity. This is particular relevant in subjects 212
with lower BAT abundance [Supplemental Figure S5 and S7]. While BAT, defined 213
with a SUV threshold of 2, was not visually apparent in these two subjects, whole fat 214
activity followed same pattern of acclimated BAT changes. When a lower SUV was 215
used (1) [Supplemental Figure S8], adipose activity changes were visually 216
concordant with overall fat activity. Mean SUV uptake in liver and skeletal muscle 217
(rectus femoris) were quantified to compare temperature-acclimation impact on BAT 218
and other metabolic organs. All images were analyzed twice by an investigator (PL) 219
blinded to subject identity and acclimation temperature. Intra-scan coefficient of 220
variation of BAT volume, mean SUV, BAT activity and mean whole fat activity were 221
0.7%, 3.0%, 1.3% and 2.4%, respectively. 222
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223
Tissue biopsies 224
Paired subcutaneous adipose/muscle biopsies were obtained at the end of each month, 225
from abdomen and rectus femoris, respectively (31). RNA extraction and cDNA 226
synthesis were performed using standard methods and genes governing thermogenesis 227
and glucose metabolism were examined using Taqman Gene Expression assays 228
(Applied Biosystems) [Supplementary Table S2]. 229
230
Laboratory measurements 231
Plasma adiponectin, leptin and fibroblast growth factor 21 (FGF21) were measured by 232
ELISA (R&D Systems, Minneapolis, MN and BioVendor, Oxford, U.K.), according 233
to manufacturer’s protocol, with intra-assay/inter-assay coefficients of variation 234
between 2.5 to 4.8%. Remaining tests were measured by Department of Laboratory 235
Medicine, NIH. 236
237
Statistical analysis 238
Statistical analysis was performed using SPSS 20.0 (SPSS, Inc., Chicago, IL, USA). 239
Data are expressed as mean±standard deviations. Trend changes of physiologic and 240
hormonal parameters during temperature acclimation across 4 months, expressed as 241
fold change over baseline, were analyzed by one-way ANOVA with Bonferroni’s 242
correction. Areas under the curve (AUC) were calculated using the trapezoidal rules 243
incorporating sampling-points across 24 hour-period from 0800 to 0700 the next 244
morning [Supplemental Figure S2]. Post-prandial glucose and insulin AUCs were 245
calculated in the period after lunch starting 1pm (T=0, 60, 120, 240, 360 minutes). 246
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Pearson correlation coefficients were used to examine associations between variables. 247
An α error of 0.05 was considered statistically significant. 248
249
RESULTS 250
251
Baseline acute thermo-metabolic evaluation 252
Five men (21±2 years old, BMI: 22±1 kg/m
2
, body fat: 21±2%) participated in the 253
study. Volunteers were first evaluated at baseline for BAT status and thermo-254
metabolic responses to temperature changes. Compared to 24°C, mild cold exposure 255
at 19°C increased total EE by 6±4% (p<0.05), representing CIT response. Baseline 256
cold-activated BAT volume was 55±61 ml with mean SUV of 3.2±0.8. EE at 19°C 257
correlated positively with BAT volume (R
2
=0.82, p=0.03). These results replicated 258
findings in our previous overnight cold exposure studies (24; 31), and validated the 259
methodology in the investigation of temperature acclimation-associated metabolic and 260
physiologic consequences. Hereafter, we describe changes in physiologic and 261
metabolic parameters at each monthly thermo-metabolic evaluation, with results 262
stratified to either 19°C or 24°C testing condition to decipher impact of acclimation 263
on metabolism under thermoneutrality and mild cold exposure. 264
265
Metabolic consequences of monthly acclimation 266
Tables 1-4 summarize changes in BAT, physiologic, dietary, body compositional and 267
hormonal parameters across 4-month acclimation. Hormone/metabolite AUC results 268
are shown in Table 4 and their fasting levels in Supplemental Table S3. Results from 269
each domain are described in the following sub-sections. 270
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271
BAT changes 272
Figure 1A-D demonstrates BAT evolution in one representative subject throughout 4-273
month sequential acclimation. Supplemental Figures S4-S7 show individual results. 274
Figure 1E-H display mean changes in BAT volume and overall fat metabolic activity, 275
which increased upon cold acclimation (19°C) by 42±18% (p<0.05) and 10±11% 276
(p<0.05), respectively; decreased after the thermoneutral month (24°C) to nearly 277
baseline level, and completely muted at the end of one-month warm exposure (27°C). 278
BAT radio-density, measured in HU, responded to acclimation with the same pattern 279
(p<0.01) [Table 1]. BAT HUs increased by 25±8% following cold acclimation, 280
reversed after the thermoneutral month, and by the end of warm acclimation in month 281
4, HU was 18±11% lower than baseline values in month 1 [Table 1]. In contrast, 282
mean SUV of skeletal muscle and liver remained unchanged during acclimation 283
[Table 1]. Room (p<0.05) and individually exposed temperatures (p<0.01), but not 284
outdoor temperatures, correlated with BAT changes during study period [Figure 1I, 1J 285
and Supplemental Table S1]. 286
287
Cold- and diet-induced thermogenesis 288
We next explored metabolic consequences of BAT acclimation. CIT response did not 289
change significantly during temperature acclimation [Table 2]. In contrast, DIT 290
measured at 19°C rose by 32±35% (p=0.03) following cold acclimation. Progressive 291
re-warming suppressed 19°C DIT response at months 3 and 4 to nearly baseline level. 292
DIT measured at 24°C was unaltered [Table 2]. 293
294
Shivering response and cold sensitivity 295
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Surface EMG recordings of muscle fasciculation/shivering measured at 19°C and 296
24°C were not different [Table 2], indicating absence of significant shivering and 297
validating our model in capturing non-shivering thermogenesis. Monthly acclimation 298
did not alter EMG recordings and subjects did not report changes in cold perception at 299
19°C during monthly calorimeter testing [Supplemental Figure S9]. 300
301
Diet and body composition 302
Neither total caloric nor macronutrient content of intake changed during acclimation 303
[Table 3]. Biweekly hunger and satiety scores did not change significantly [Table 3]; 304
however, volunteers reported an increase in desire to eat and reduction in satiety 305
during ad libitum meal test after cold acclimation, which reversed during the warm 306
months [Supplemental Figure S10]. Body composition was unaltered across study 307
period [Table 3]. 308
309
Pituitary-thyroid-adrenal axis 310
To elucidate potential endocrine mediators of BAT acclimation, we profiled pituitary-311
thyroid-adrenal axes [Table 4]. Cold acclimation increased free triiodothyronine (T3) 312
AUC measured at 24°C, but not at 19°C. Free T3 to free thyroxine (T4) ratio (an 313
indicator of peripheral T4 to T3 conversion (34)) was greater by 11±5% (p=0.01) 314
measured at 24°C. No significant changes were observed in TSH or the pituitary-315
adrenal axis. 316
317
Insulin sensitivity 318
Total glucose and insulin AUCs did not change during acclimation [Table 4]. In 319
contrast, post-prandial insulin excursion measured at 19°C reached a nadir after cold 320
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acclimation, without significant changes to glucose excursion [Figure 2A-B]. Indices 321
of insulin sensitivity and resistance showed significant reciprocal changes during 322
cold- and warm-acclimation, consistent with an improvement of post-prandial whole 323
body insulin sensitivity following cold acclimation [Figure 2C-D]. These changes 324
were absent during measurements at 24°C [Figure 3A-D]. 325
326
Adipokine changes 327
Given our recent demonstration of BAT as an endocrine organ in humans (19; 32; 328
35), we probed adipokine changes during acclimation. Adiponectin AUC was 329
augmented by 22±9% (p<0.001) after cold acclimation [Figure 2E]. Enhancement of 330
adiponectin levels was observed not only at 19°C during acute thermo-metabolic 331
evaluation, but similar increase occurred also at 24°C (p<0.001) [Figure 3E]. In 332
contrast, cold acclimation reduced leptin AUC by 14±28% (p<0.001), evident at both 333
19°C [Figure 2F] and 24°C [Figure 3F]. These dichotomized changes returned almost 334
to baseline during the thermoneutral third month, trending to the opposite directions at 335
the end of the fourth month at 27°C (p<0.05). Changes in circulating adiponectin and 336
leptin correlated negatively with changes in BAT activity after cold acclimation 337
[Figure 2G-H and Figure 3G-H]. FGF21 AUC rose after cold acclimation, although 338
overall trend did not reach significance [Table 4]. 339
340
Fat and muscle gene expression 341
To explore sources of adipokine and origins of metabolic changes, fat and muscle 342
biopsies were obtained from 4 volunteers at the end of each month. Adiponectin and 343
GLUT4 expression in adipose tissue [Figure 2I], but not muscle [Figure 3I], rose after 344
cold acclimation while expression of leptin fell, and their respective trends reversed 345
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after thermoneutral and warm acclimation months (p<0.05). Expression of CIDEA, a 346
BAT gene governing lipid mobilization (36), increased following cold acclimation but 347
decreased during re-warming [Figure 4]. No other BAT/beige fat gene changes were 348
observed. 349
350
351
Discussion 352
353
The major finding of our study is the demonstration of BAT acclimation and its 354
metabolic consequences by minimal manipulation of overnight temperature exposure, 355
while allowing usual daily activities. Human BAT is inducible and suppressible by 356
controlled mild cold and warm exposure, respectively, independent of seasonal 357
fluctuations. BAT acclimation is accompanied by boosting of diet-induced 358
thermogenesis and post-prandial insulin sensitivity. Mechanistically, this is associated 359
with reciprocal changes of circulating adiponectin and leptin, mirrored by 360
corresponding transcriptosomal changes in adipose tissue ex vivo. These results 361
provide first evidence linking ambient temperature, BAT acclimation and whole body 362
energy/substrate metabolism in humans. 363
364
Consistent with previous reports (25; 26), we confirmed BAT recruitability by cold 365
exposure, but did not observe significant CIT response augmentation; the latter could 366
be a type 2 error. Despite tentalizing associative data linking BAT abundance with 367
favorable energy metabolism in humans, it remains unclear, to date, whether BAT 368
recruitment is accompanied by metabolic benefits. We specifically sought to 369
determine the significance of BAT recruitment, and revealed an association of BAT 370
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acclimation with enhancement of post-prandial energy metabolism and insulin 371
sensitization. Within the allowance and feasibility of human research, we explored 372
underlying mechanisms through blood and tissue analyses. 373
374
First, within the pituitary-thyroid-adrenal axis, we observed an increase in T3/T4 375
ratio, which indicates enhanced T3 synthesis. Given the enrichment of BAT with type 376
2 deiodinase (37), and our previous report showing severe insulin resistance 377
amelioration by thyroid hormone-mediated BAT activation (38), we hypothesize 378
heightened T3 synthesis within BAT to be one plausible mechanism underlying 379
acclimated-BAT associated metabolic changes. Such pattern of increased thyroid 380
hormone turnover in the absence of TSH changes is reminiscent of cold adaptation 381
observed among Arctic residents (39). 382
383
Second, our adipokine profiling uncovered an intriguing relation between BAT, 384
adiponectin and leptin. Cold acclimation augmented circulating adiponectin but 385
decreased leptin. It is tempting to speculate cold-induced adiponectin, a potent 386
insulin-sensitizer, contributes to glucose metabolism improvement and leptin 387
reduction, the latter as a result of improved tissue sensitivity. Concordant gene 388
changes in adipose adiponectin and leptin, absent in muscle, argue adipose to be the 389
primary effector. Surprisingly, circulating adiponectin related negatively with BAT 390
activity, suggesting PET-detectable BAT was not the source of cold-induced 391
adiponectin. As BAT exhibits insulin-independent glucose uptake capacity (40), 392
lesser BAT expansion could have triggered alternative glucose utilizing pathways in 393
WAT during cold acclimation, evident by observed WAT GLUT4 up-regulation. 394
Interestingly, such changes in circulating adiponectin and leptin were not limited to 395
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cold-exposed condition [Figure 2], but persisted at thermoneutrality [Figure 3], 396
indicating the temperature-acclimated hormonal milieu was not totally dependent on 397
BAT activation. The corollary is that acclimated BAT could be serving beneficial 398
metabolic functions not related to temperature regulation per se. 399
400
Third, newly identified cytokines, such as FGF21, may mediate temperature-401
acclimated tissue crosstalk. Recent identification of a FGF21-adiponectin feed-402
forward axis (41) led us to wonder if FGF21 augmentation following cold-acclimation 403
could have brought forth the adiponectin rise. When BAT was muted at the end of 404
warm acclimation, and adiponectin dwindled, FGF21 did not fall however, suggesting 405
non-BAT FGF-secreting tissues might have compensated in states of relative BAT 406
deficiency. 407
408
Fourth, although we did not observe an increase in beige fat gene expression, possibly 409
due to the small sample size, we speculate fat browning to be a possibility. This is 410
corroborated by finding an increased expression of the BAT gene CIDEA in adipose 411
tissue following cold acclimation. Although ethical considerations prohibited serial 412
neck fat biopsies in our volunteers, changes in radio-density within BAT by PET/CT 413
have offered insight on tissue changes. Adipose tissue is typically characterized by 414
HU between -10 to -300, in contrast to muscle tissue, whose HU is within the positive 415
range. Compared to WAT, BAT has relatively less lipid, as it is filled with abundant 416
mitochondria and blood vessels. This is exemplified by water-fat separated magnetic 417
resonance imaging revealing lower fat fraction in activated BAT both in humans (42) 418
and rodents (43). We speculate the rise and fall in BAT radio-density with cold and 419
warm acclimation, respectively, could be reflections of WATBAT transformation 420
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(or fat brown-ing). This is also supported by previous studies demonstrating cell-421
autonomous (44) and endocrine-mediated (19) cold-induced WAT browning in 422
humans. Further studies are required to ascertain if WAT browning contributes to 423
cold-acclimated BAT induced metabolic changes. 424
425
Collectively, our results infer a complex concerted BAT-WAT response to cold 426
acclimation, which could involve interplay between CIDEA-mediated lipid 427
mobilization (45; 46), GLUT4-enhanced glucose utilization and FGF21/adiponectin-428
induced insulin sensitization. Most importantly, all these changes occurred in the 429
absence of measureable EE, caloric intake or body compositional alterations, 430
suggesting such responses to be primary cold-induced metabolic sequelae, rather than 431
compensatory physiologic adaptations. Nonetheless, because the desire to eat 432
heightened after cold acclimation, we cannot exclude the possibility that appetite 433
stimulation could diminish metabolic benefits of BAT recruitment if it increases 434
caloric intake in longer-term studies. 435
436
The inducibility, suppressibility and plasticity of human BAT entail implications 437
beyond thermoregulatory physiology. The translation of recently discovered BAT-438
activators in the laboratory to pharmacologic BAT stimulants available for clinical 439
use is not a trivial process (23). Our study substantiates, in contrast, a simple BAT-440
modulating strategy: a mild reduction in environmental temperature is capable in 441
recruiting BAT and yielding associated metabolic benefits; conversely, even a small 442
elevation in ambient temperature could impair BAT, and dampen previously attained 443
metabolic benefits. Such reversible metabolic switching, occurring within a 444
temperature range achievable in climate-controlled buildings, therefore carries 445
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therapeutic implications of BAT-acclimation, both on an individual and a public 446
health level. Bedroom temperature has gradually increased from 19°C to 21.5°C over 447
the last 3 decades in the US (47). The blunting of BAT function due to widespread 448
use of indoor climate control could be a neglected contribution to the obesity 449
epidemic. Moderate downward adjustment of indoor temperature could represent a 450
simple and plausible strategy in dampening the escalation of obesity on a population 451
level. Our volunteers reported satisfactory sleep during acclimation, although more 452
formal assessment of sleep quality is required in future studies. 453
454
Our findings should be viewed as a proof of concept illustrating human BAT 455
plasticity. We acknowledge the small sample size to be a limitation of our study. 456
Unfortunately, the conduct of long-term acclimation study necessitated substantial 457
resources and regrettably prohibited a large sample size. Despite a small study 458
population, the investigations were undertaken in a tightly monitored and controlled, 459
yet real life-simulating and applicable setting, encompassing the most comprehensive 460
spectrum of energy balance/metabolism to date to tackle a question fundamental to 461
human BAT research: what is the significance of BAT recruitment? The unveiled 462
positive relation between acclimated-BAT and glucose homeostasis is clinically 463
relevant. Glucose intolerance is an independent risk factor of cardiovascular mortality 464
and post-prandial hyperglycemia is its earliest manifestation (48). We emphasize a 465
causal linkage could not be definitely ascertained between BAT recruitment and post-466
prandial insulin sensitivity improvement; however our study provides compelling 467
circumstantial evidence supporting a potential therapeutic role of BAT in impaired 468
glucose metabolism, and calls for the investigation of similar temperature acclimation 469
in individuals with impaired glycemia. Our observation of BAT recruitment 470
Page 19 of 59 Diabetes
20
accompanied by insulin sensitization in the absence of significant weight loss echoes 471
animal findings showing glucose homeostasis improvement following fat browning to 472
be greater than expected from adiposity reduction alone (49). Whether it was indeed a 473
result of fat phenotypic and/or adipokine changes merits further studies. 474
475
In summary, temperature acclimation modulates BAT abundance and activity, 476
subsequently impacting energy and substrate metabolism in humans. BAT exhibits 477
thermal plasticity intimately related to glucose homeostasis. Harnessing BAT by 478
simple adjustment of ambient temperature could be a new strategy in the combat 479
against obesity, diabetes and related disorders. 480
481
Page 20 of 59Diabetes
21
Acknowledgements 482
483
Paul Lee was supported by an Australian National Health Medical Research Council 484
(NHMRC) Early Career Fellowship, the Diabetes Australia Fellowship and Bushell 485
Travelling Fellowship, and the School of Medicine, University of Queensland. This 486
study was supported by the Intramural Research Program Z01-DK047057-07 of 487
NIDDK and the NIH Clinical Center. We thank Dr Peter Herscovitch and Dr Corina 488
Millo, both from PET Department, Clinical Center, NIH, for advice on PET-CT 489
scanning; Rachel Perron, Christopher Idelson, Sarah Smyth, Jacob Hattenbach and 490
Juan Wang, all from Diabetes Endocrinology Obesity Branch, NIDDK, NIH, for 491
technical assistance; Dilalat Bello and Oretha Potts, from Clinical Center, NIH, for 492
dietary counseling/monitoring, and all nurses in the Clinical Metabolic Unit, NIH, for 493
their nursing care. 494
495
P.L., S.S., J.L., A.B.C., R.J.B., K.Y.C., and F.S.C. participated in study concept, 496
design, research, acquisition of data, analysis and discussion of results. W.D. and 497
C.D.W. researched and anslyzed data, and contributed to discussion of results. P.L. 498
wrote the article, and all authors participated in critical revision and approved the final 499
version of the manuscript. 500
501
P.L. and F.S.C. are the guarantors of this work and, as such, had full access to all of 502
the data in the study and take responsibility for the integrity of the data and the 503
accuracy of the data analysis. 504
505
Page 21 of 59 Diabetes
22
The funders have no role in the design and conduct of the study; collection, 506
management, analysis, and interpretation of the data; and preparation, review, or 507
approval of the manuscript. 508
509
No potential conflicts of interest relevant to this article were reported. 510
511
512
513
514
515
516
517
518
519
520
Page 22 of 59Diabetes
23
Figure legends 521
522
Figure 1 Temperature-dependent BAT acclimation Panels A-D display 523
representative PET-CT fused images of the cervical-supraclavicular region (left panel: 524
coronal view; right panel: transverse view) of one subject during monthly temperature 525
acclimation. BAT (Hounsfield units: -300 to -10 and SUV2) was shown in red. 526
Baseline BAT volume, mean SUV and activity were 26 ml, 2.65 and 0.238 MBq, 527
respectively [Panel A]. All parameters increased following one month of mild cold 528
acclimation (19°C) [Panel B], decreased to nearly baseline level after thermoneutral 529
month (24°C) [Panel C], and was nearly completely abolished at the end of 1-month 530
mild warm exposure in the final month (27°C) [Panel D]. Mean fold changes (N=5) of 531
BAT volume [Panel E], mean SUV [Panel F] and BAT activity [Panel G], relative to 532
month 1 (24°C), were significant across 4-month acclimation. Whole fat activity, as 533
defined by
18
F-fluodeoxyglucose uptake within tissue of fat density (Hounsfield units: 534
-300 to -10), followed the same pattern [Panel H], and interacted significantly with 535
temperature acclimation. Room [Panel I] and individual exposed temperatures [Panel 536
J], but not environmental seasonal fluctuations [Panel I], tracked BAT and whole fat 537
metabolic changes in the predicted temperature-dependent manner. Correlative 538
analysis between BAT parameters and temperature exposure is shown in 539
Supplemental Table S1. Individual PET-CT images and temperature profiles are 540
shown in Supplemental Figures S4-S7. *p<0.05 compared to month 1 (24°C); 541
#p<0.05 compared to month 2 (19°C). 542
543
544
Page 23 of 59 Diabetes
24
Figure 2 Metabolic consequences of BAT-acclimation at 19°
°°
°C Panels A and B 545
compare post-prandial glucose and insulin excursions after mixed meal at 13:00 546
before and after cold acclimation, respectively, measured at 19°C. Glucose excursions 547
were unchanged but insulin levels decreased, with a significant reduction in AUC, 548
following mild cold acclimation (month 2). Accordingly, adipocyte insulin resistance 549
(IR) was the lowest [Panel C], and Matsuda index (an indicator of insulin sensitivity) 550
was the highest [Panel D] after cold acclimation (month 2). These changes in glucose 551
metabolism were accompanied by an increase in circulating adiponectin [Panel E] and 552
a decrease in circulating leptin [Panel F]. Cold acclimation-induced changes (month 1 553
to 2) in circulating adiponectin [Panel G] and leptin levels [Panel H] correlated 554
negatively with changes in BAT activity. Adiponectin and leptin mRNA displayed 555
concordant changes in subcutaneous adipose tissue biopsies with circulating levels 556
and changes in GLUT4 tracked those of adiponectin [Panel I].
a
p<0.05 compared to 557
month 1 (24°C),
b
p<0.05 compared to month 2 (19°C),
c
p<0.05 compared to month 3 558
(24°C) and
d
p<0.05 compared to month 4 (27°C). 559
560
Figure 3 Metabolic consequences of BAT-acclimatization at 24°
°°
°C. Panels A and B 561
compare post-prandial glucose and insulin excursions after mixed meal at 13:00 562
before and after cold acclimatization, respectively, measured at 24°C. Unlike 563
measurements at 19°C [Figure 2A and B], no significant changes were observed in 564
glucose or insulin excursions. Accordingly, adipocyte insulin resistance (IR) [Panel 565
C] and Matsuda index (an indicator of insulin sensitivity) [Panel D] were unchanged. 566
However, Circulating adiponectin increased [Panel E], while leptin decreased [Panel
567
F], identical to measurements observed at 19°C [Figure 2E and F]. Cold 568
acclimatization-induced changes (month 1 to 2) in circulating adiponectin [Panel G] 569
Page 24 of 59Diabetes
25
and leptin levels [Panel H] correlated negatively with changes in BAT activity. In 570
contrast to those observed in adipose tissue [Figure 2I], Adiponectin and GLUT4 571
mRNA did not change significantly in muscle [Panel I].
c
p<0.05 compared to month 3 572
(24°C) and
d
p<0.05 compared to month 4 (27°C). 573
574
Figure 4 BAT and beige fat gene changes in adipose tissue biopsies across 4-575
month acclimatization. Panel A shows changes in general BAT gene expression 576
(general BAT genes are defined as genes ascribed to general BAT function, and do 577
not indicate their developmental origin). Expression of CIDEA, but not others, 578
changed significantly (p=0.04) during acclimatization across 4-month period. Panel B 579
shows changes in classic BAT gene expression. Classic BAT genes are defined as 580
those expressed in interscapular BAT in animals or human infants (50). Panel C 581
showed changes in beige fat gene expression. Beige fat genes are defined as those 582
expressed in inducible brown adipocytes, also known as brite or beige adipocytes, 583
found within WAT depots. No significant changes were observed in classic BAT 584
and beige fat genes across temperature acclimation. 585
586
587
588
Page 25 of 59 Diabetes
26
Table 1 PET-CT parameters across 4 months of acclimation At the end of each 589
testing month, subjects underwent acute thermo-metabolic evaluation at either 24°C 590
or 19°C. Results are reported as mean±standard deviation.
a
p<0.05 (month 1 vs. 2). 591
592
Month 1
24°
°°
°C
Month 2
19°
°°
°C
Month 3
24°
°°
°C
Month 4
27°
°°
°C
Trend
P value
PET-CT parameters
BAT volume
(ml)
55
±
61 78
±
84
a
63
±
81 58
±
81 0.036
BAT mean
SUV
3.2
±
0.8 3.8
±
1.3 3.4
±
1.0 3.4
±
0.8 0.35
BAT activity
(MBq)
0.65
±
0.76 1.0
±
1.3
a
0.8
±
1.1 0.7
±
1.0 0.038
BAT
radiodensity
(Houndsfield
units)
-58.8
±
7.2 -44.2
±
6.8 -55.4
±
6.5 -69.2
±
6.8 <0.01
Whole fat
mean SUV
0.61
±
0.13 0.68
±
0.18
a
0.63
±
0.17 0.59
±
0.16 0.035
Muscle mean
SUV
0.46
±
0.08 0.41
±
0.04 0.43
±
0.05 0.48
±
0.08 0.52
Liver mean
SUV
1.68
±
0.08 1.50
±
0.14 1.61
±
0.15 1.67
±
0.16 0.15
Page 26 of 59Diabetes
27
Table 2 Physiologic parameters across 4 months of acclimation At the end of each testing month, subjects underwent acute thermo-metabolic 593
evaluation at either 24°C or 19°C. Results are reported as mean±standard deviation.
a
p<0.05 compared to 24°C during acute thermo-metabolic 594
evaluation each month;
b
p<0.05 (month 1 vs. 2), compared to matching measurement at same temperature performed at respective months as 595
indicated. 596
597
Month 1
24°
°°
°C
Month 2
19°
°°
°C
Month 3
24°
°°
°C
Month 4
27°
°°
°C
Trend
P value
Physiologic parameters
Calorimeter
°
C
24
°
C
19
°
C
24
°
C
19
°
C
24
°
C
19
°
C
24
°
C
19
°
C
24
°
C
19
°
C
Total EE (kcal) 2472
±
180 2624
±
198
a
2366
±
358 2543
±
410
a
2400
±
252 2555
±
346
a
2341
±
255 2505
±
322
a
0.45 0.46
Respiratory
quotient
0.84
±
0.03 0.84
±
0.01 0.84
±
0.02 0.83
±
0.02 0.85
±
0.02 0.84
±
0.02 0.84v0.03 0.85
±
0.02 0.72 0.47
Total activity
(units)
8.4
±
1.6 8.4
±
2.8 7.6
±
2.9 7.1
±
2.4 6.8
±
3.7 7.1
±
4.1 7.9
±
5.0 7.2
±
4.7 0.54 0.38
Surface
electromyography
(x10
-6
RMS)
2.8
±
0.4 2.5
±
1.3 2.7
±
0.3 2.6
±
0.2 2.8
±
0.5 2.7
±
0.2 2.8
±
0.3 2.6
±
0.4 0.98 0.83
CIT (%)
6.2
±
4.1
7.4
±
3.1
6.2
±
3.9
6.8
±
3.2
0.16
DIT (%) 10.3
±
13.1 33.4
±
18.2 19.0
±
15.4 42.2
±
17.4
a,b
19.1
±
16.0 37.1
±
19.6
a
22.5
±
11.0 34.4
±
19.2
a
0.30 0.36
598
599
Page 27 of 59 Diabetes
28
Table 3 Nutritional and body compositional parameters across 4 months of acclimation At the end of each testing month, subjects 600
underwent acute thermo-metabolic evaluation at either 24°C or 19°C. Results are reported as mean±standard deviation.
a
p<0.05 (month 1 vs. 2), 601
b
p<0.05 (months 2 vs. 4), compared to matching measurement at same temperature performed at respective months as indicated. 602
603
Month 1
24°
°°
°C
Month 2
19°
°°
°C
Month 3
24°
°°
°C
Month 4
27°
°°
°C
Trend
P value
Dietary intake
Caloric (kcal/d)
2530
±
321
2620
±
412
2623
±
342
2514
±
359
0.32
Protein (g/d)
126
±
16
131
±
19
131
±
15
127
±
18
0.35
Fat (g/d)
88
±
9
91
±
13
93
±
11
86
±
10
0.16
Carbohydrate
(g/d)
319
±
42 331
±
51 329
±
44 320
±
48 0.44
Appetite/hunger visual analogue scale
Hunger AUC 13.1
±
6.3 20.7
±
7.0 17.5
±
6.1 15.5
±
3.5 0.13
Satiety AUC 35.8
±
4.6 25.7
±
8.5 25.5
±
5.4 27.9
±
7.9 0.09
Desire to eat AUC 14.6
±
4.3 20.1
±
3.4
a,b
17.9
±
4.2 16.5
±
5.3 0.003
Body composition
Body weight (kg)
74.4
±
7.3
74.8
±
7.5
74.9
±
7.4
74.7
±
7.7
0.72
Lean mass (kg)
55.8
±
6.0
56.3
±
6.1
56.6
±
6.3
56.1
±
6.4
0.56
Fat mass (kg)
14.6
±
0.5
14.5
±
0.8
14.6
±
1.4
14.7
±
1.7
0.95
% body fat
20.92
±
2.00
20.62
±
1.64
20.64
±
2.22
20.88
±
2.51
0.99
604
605
Page 28 of 59Diabetes
29
Table 4 Hormonal and metabolic parameters across 4 months of acclimation At the end of each testing month, subjects underwent acute 606
thermo-metabolic evaluation at either 24°C or 19°C. Results are reported as mean±standard deviation.
a
p<0.05 compared to 24°C during acute 607
thermo-metabolic evaluation each month;
b
p<0.05 (month 1 vs. 2),
c
p<0.05 (months 2 vs. 4) and
d
p<0.05 (months 1 vs. 4), compared to matching 608
measurement at same temperature performed at respective months as indicated. 609
610
Month 1
24°
°°
°C
Month 2
19°
°°
°C
Month 3
24°
°°
°C
Month 4
27°
°°
°C
Trend
P value
Hormonal parameters
Calorimeter
°
C
24
°
C
19
°
C
24
°
C
19
°
C
24
°
C
19
°
C
24
°
C
19
°
C
24
°
C
19
°
C
Sympathoadrenal
Urinary
epinephrine (ug/d)
7.5
±
5.0 8.0
±
4.8 7.5
±
3.9 8.3
±
5.6 7.5
±
4.4 7.7
±
4.6 8.3
±
6.1 7.7
±
5.1 0.97 0.94
Urinary
norepinephrine
(ug/d)
46
±
29 53
±
14 56
±
30 64
±
20 35
±
9 61
±
18
a
37
±
5 58
±
24 0.42 0.56
Glucocorticoid axis
ACTH AUC
(pg.min/ml)
207
±
61 199
±
73 199
±
51 197
±
65 203
±
74 176
±
61 209
±
71 199
±
83 0.20 0.40
Cortisol AUC
(µg.min/ml)
0.96
±
0.14 0.91
±
0.13 0.87
±
0.10 0.95
±
0.12 0.81
±
0.13 0.84
±
0.17 0.90
±
0.15 0.88
±
0.22 0.14 0.39
Urinary cortisol
(µg/d)
49
±
9 36
±
13 49
±
27 39
±
9 39
±
11 38
±
11 39
±
22 49
±
21 0.42 0.25
Thyroid axis
TSH AUC
(µIU.min/ml)
7.8
±
3.5 8.0
±
2.8 7.5
±
3.6 7.3
±
2.1 8.6
±
4.3 7.3
±
2.2 8.8
±
3.5
c
7.6
±
1.9 0.38 0.39
Page 29 of 59 Diabetes
30
Free T4 AUC
(pg.min/ml)
95
±
13 95
±
9 93
±
12 92
±
12 93
±
9 93
±
11 90
±
9 94
±
9 0.39 0.54
Free T3 AUC
(pg.min/ml)
26
±
1 27
±
1 29
±
2
b
28
±
1 28
±
3 27
±
2 26
±
2
d
28
±
2 0.16 0.59
Free T3/free T4
AUC
2381
±
490 2410
±
345 2642
±
574
b
2556
±
393 2513
±
449 2511
±
466 2515
±
481 2491
±
403 0.06 0.41
Glucose and lipid metabolism
Total glucose
AUC (mg.min/ml)
7.38
±
0.64 7.32
±
0.37 7.22
±
0.51 7.25
±
0.63 7.22
±
0.73 7.32
±
0.63 7.14
±
0.51 7.11
±
0.74 0.75 0.71
Post prandial
glucose AUC
(mg.min/ml)
2.73
±
0.27 2.68
±
0.34 2.59
±
0.13 2.68
±
0.29 2.64
±
0.40 2.57
±
0.29 2.64
±
0.15 2.53
±
0.27 0.79 0.63
Total insulin AUC
(IU.min/L)
170
±
102 210
±
83 198
±
97 143
±
49 212
±
118 186
±
123 171
±
87 182
±
128 0.08 0.44
Post prandial
insulin AUC
(IU.min/L)
106
±
64 133
±
57 111
±
52 77
±
22 132
±
85 114
±
80 103
±
55 109
±
79 0.19 0.31
Total free fatty
acid AUC
(mEq.min/L)
3.53
±
0.70 3.76
±
1.05 3.36
±
0.31 3.37
±
1.16 2.73
±
0.86 3.20
±
0.40 3.65
±
1.09 3.76
±
0.41 0.36 0.68
Fasting total
cholesterol
(mg/dL)
120
±
24 132
±
24 117
±
16 136
±
11 0.13
Fasting LDL
(mg/dL)
71
±
21 75
±
21 62
±
13 76
±
10 0.20
Fasting TG
(mg/dL)
57
±
16 68
±
27 68
±
15 65
±
22 0.31
Fasting HDL
(mg/dL)
38
±
7 44
±
7 41
±
7 46
±
4
d
0.03
Adipokine
Leptin AUC
(ng.min/ml)
16
±
5 15
±
6 14
±
3
b
12
±
2
a,b
29
±
15 26
±
11 25
±
12 25
±
11
c
0.01 0.002
Page 30 of 59Diabetes
31
Adiponectin AUC
(pg.min/ml)
99
±
38 103
±
37 117
±
51
b
127
±
49
a,b
78
±
31 77
±
32 74
±
32
c
82
±
38
c
0.0007 0.0003
FGF21 AUC
(pg.min/ml)
333
±
57 411
±
104 343
±
46 460
±
91
a,b
350
±
25 400
±
80 370
±
37 435
±
75 0.28 0.10
611
612
613
614
615
616
Page 31 of 59 Diabetes
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Temperature-dependent BAT acclimation
254x338mm (72 x 72 DPI)
Page 36 of 59Diabetes
Metabolic consequences of BAT-acclimation at 19°C
254x190mm (72 x 72 DPI)
Page 37 of 59 Diabetes
Metabolic consequences of BAT-acclimatization at 24°C
254x190mm (72 x 72 DPI)
Page 38 of 59Diabetes
BAT and beige fat gene changes in adipose tissue biopsies across 4-month acclimatization
254x338mm (72 x 72 DPI)
Page 39 of 59 Diabetes
1
SUPPLEMENTAL DATA 1
2
Supplement to: Lee et al. Temperature-acclimated brown adipose tissue modulates 3
insulin sensitivity in humans 4
5
6
Contents 7
Table S1-S3 8
Figure S1-S10 9
References 10
11
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2
Table S1 Correlation coefficients between BAT parameter fold changes and temperature exposure over 4-month 12
acclimatization. No relationships were seen between BAT parameters and environmental seasonal fluctuations. In contrast, strong 13
correlations were observed between controlled room temperature and individual exposed temperatures with BAT and whole fat 14
metabolic activity. *p<0.05, #p<0.01 and ^^p<0.10. 15
16
Pearson Correlation coefficients BAT
volume
Mean
SUV
BAT
activity
Whole
fat
activity
Environmental
Temp
Room
Temp
Outside clothing
Temp
Under clothing
Temp
Max Min Day Night Day Night Day Night
BAT volume 0.56 0.99* 0.97* 0.63 0.76 -0.98* -0.98* -1.00
#
-0.99* -0.98* -0.85
Mean SUV 0.56 0.67 0.68 0.02 0.06 -0.48 -0.47 -0.57 -0.50 -0.69 -0.66
BAT activity 0.99* 0.67 0.99
#
0.60 0.71 -0.97* -0.97* -0.99* -0.98* -1.00
#
-0.89
Whole fat activity 0.97* 0.68 0.99
#
0.65 0.74 -0.97* -0.97* -0.97* -0.97* -0.99
#
-0.94
^^
Environmental maximum temp 0.63 0.02 0.60 0.65 0.97* -0.77 -0.77 -0.60 -0.73 -0.58 -0.76
Environmental minimum temp 0.76 0.06 0.71 0.74 0.97* -0.87 -0.87 -0.74 -0.84 -0.70 -0.78
Room day temp (day) -0.98* -0.48 -0.97* -0.97* -0.77 -0.87 1.00
#
0.97* 1.00
#
0.96* 0.90
^^
Room temp (night) -0.98* -0.47 -0.97* -0.97* -0.77 -0.87 1.00
#
0.97* 1.00
#
0.96* 0.89
Outside clothing temp (day) -1.00
#
-0.57 -0.99* -0.97* -0.60 -0.74 0.97* 0.97* 0.98* 0.98* 0.84
Outside clothing temp (night) -0.99* -0.50 -0.98* -0.97* -0.73 -0.84 1.00
#
1.00
#
0.98* 0.97* 0.89
Under clothing temp (day) -0.98* -0.69 -1.00
#
-0.99
#
-0.58 -0.70 0.96* 0.96* 0.98* 0.97* 0.90
^^
Under clothing temp (night) -0.85 -0.66 -0.89 -0.94
^^
-0.76 -0.78 0.90
^^
0.89 0.84 0.89 0.90
^^
17
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3
Table S2 Taqman gene expression assays 18
19
Gene name Catalogue number
Adiponectin Hs00605917_m1
Leptin Hs00174497_m1
GLUT4 Hs00168966_m1
UCP1 Hs00222452_m1
PRDM16 Hs00922674_m1
PGC1
α
αα
α
Hs01016719_m1
CIDEA Hs00154455_m1
DIO2 Hs00988260_m1
ZIC1 Hs00602749_m1
LHX8 Hs00418293_m1
TBX1 Hs00271949_m1
TMEM26 Hs00415619_m1
HOXC9 Hs00396786_m1
FGF21 Hs00173927_m1
TBP Hs00427620_m1
20
21
22
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4
Table S3 Fasting hormonal and metabolic parameters across 4 months of acclimation At the end of each testing month, subjects 23
underwent acute thermo-metabolic evaluation at either 24°C or 19°C. Results of measurements obtained at 7am after 24 hour exposure 24
to either 24°C or 19°C are reported as mean±standard deviation.
a
p<0.05 compared to 24°C during acute thermo-metabolic evaluation 25
each month;
b
p<0.05 (month 1 vs. 2),
c
p<0.05 (months 2 vs. 4),
d
p<0.05 (months 1 vs. 4), and
e
p<0.05 (months 2 vs. 3), compared to 26
matching measurement at same temperature performed at respective months as indicated. 27
28
Month 1
24°
°°
°C
Month 2
19°
°°
°C
Month 3
24°
°°
°C
Month 4
27°
°°
°C
Trend
P value
Hormonal parameters
Calorimeter
°
C 24
°
C 19
°
C 24
°
C 19
°
C 24
°
C 19
°
C 24
°
C 19
°
C 24
°
C 19
°
C
Glucocorticoid axis
ACTH
(pg/ml)
25.2±9.9 29.4±13.6 35.0±16.4 28.0±8.4 24.8±10.2 29.6±13.4 28.1±17.4 23.8±11.9 0.60 0.21
Cortisol
(µg/dl)
15.5±2.7 15.0±4.1 16.7±4.6 15.9±5.0 14.0±1.9 14.9±5.6 12.7±1.6 12.2±3.9 0.26 0.53
Thyroid axis
TSH
(µIU/ml)
1.1±0.5 1.3±0.5 1.1±0.6 1.3±0.5 1.3±0.6 1.5±0.7 1.3±0.6 1.3±0.4 0.29 0.59
Free T4
(ng/dL)
1.1±0.1 1.1±0.1 1.1±0.1 1.1±0.2 1.1±0.1 1.1±0.1 1.1±0.1 1.1±0.1 0.74 0.93
Free T3
(µg/dl)
312±29 334±12 340±11 337±14 313±6 346±49 341±48 328±24 0.14 0.66
Glucose and lipid metabolism
Glucose
(mg/ml)
85.8±4.2 85.2±3.7 82.6±6.4 85.0±5.7 90.0±6.0 84.4±16.3 84.6±6.1 86.8±3.3 0.19 0.98
Page 43 of 59 Diabetes
5
Insulin
(IU/L)
6.2±3.5 8.4±2.5 10.6±5.0 7.6±2.6 7.6±3.0 8.4±4.0 8.2±3.9 8.4±4.0 0.26 0.95
Free fatty acid
(mEq/L)
0.3±0.2 0.3±0.1 0.4±0.1 0.3±0.1 0.3±0.1 0.2±0.1 0.4±0.2 0.3±0.1 0.35 0.22
Adipokine
Leptin
(ng/ml)
2.7±1.1
d
2.6±1.1
d
2.2±0.8
c
1.9±0.6
c
,e
4.6±2.1 4.1±1.8
a
4.8±2.1 4.1±1.8
a
<0.01 <0.01
Adiponectin
(pg/ml)
10.6±4.1
10.6±4.2 12.7±5.4 13.7±5.8
a
,b,c
,e
8.0±3.3 8.2±3.5 8.0±3.8 8.5±3.9 <0.01 <0.01
29
30
31
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6
Figure legends 32
33
Figure S1 Flow chart. Flow chart of volunteer recruitment, allocation and intervention. 34
35
Figure S2 Overall outline of acclimatization and thermo-metabolic evaluation 36
protocol This is a prospective crossover study consisting of 4 consecutive blocks of 37
studies of 1-month duration each. It incorporates i) sequential monthly thermal 38
acclimatization over a 4-month period, and ii) acute thermo-metabolic evaluations at the 39
end of each study temperature regime. Procedures undertaken during each acute thermo-40
metabolic evaluation were listed. Volunteers underwent two 24-hour sessions of 41
observation, while exposed to first 24°C and then 19°C, with a resting non-testing period 42
of 1 day in between. PET-CT scanning was performed after the 19°C testing day at the 43
end of each acclimatization month. Each subject undertook a total of four PET-CT scans 44
during the entire study. During the two 24-hour sessions, volunteers wore standardized 45
hospital clothing with a combined thermal insulation value of 0.4 (clo). Subjects were 46
fasted in the morning (8 hours from previous night) to allow fasting blood samples to be 47
collected. The meals served during study were caffeine-free with fixed macronutrient 48
contents (Lunch and dinner: one-third and two-thirds of daily caloric intake, 49
respectively). Volunteers were informed to minimize physical activity during testing. 50
Hormonal and metabolic parameters were measured in the calorimeter, at time points as 51
indicated, to allow AUC computation (FGF21 AUC was calculated incorporating 5 time 52
points (0, 1, 4, 5, 9 hours); AUC of other hormones/substrates incorporated all 10 time 53
points.) The following procedures were undertaken during the two 24-hour thermo-54
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7
metabolic evaluations: real-time energy expenditure, RQ, spontaneous movements, blood 55
sampling for hormonal/substrate measurements, 12-hour urine collection for 56
catecholamine and cortisol, and optional subcutaneous adipose tissue and/or muscle 57
biopsy. 58
59
Figure S3 Three-dimensional region of interest (ROI) constructed for comparison of 60
BAT volume and activity across 4-month period. The region was defined cranially by 61
a horizontal line (blue) parallel to the base of C4 vertebra, and caudally by an oblique line 62
(maroon) traversing the manubriosternal joint and superior aspect of the T8 transverse 63
process. Panel A displayed sagittal sections of PET-CT of one subject, showing the 64
defined ROI extending from midline to the arm (left to right). Coronal sections are 65
displayed, extending anteriorly from sternoclavicular joint to the vertebral column 66
posteriorly in Panel B (left to right). The ROI captures all major BAT depots (in red) in 67
humans: cervical, supraclavicular, superior mediastinal, axillary and paravertebral depots, 68
as shown in the sagittal and coronal images. 69
70
Figure S4-S7 Temperature-dependent BAT acclimatization of individual subjects 71
Panels A-D in each Figure show representative PET-CT fused images of the cervical-72
supraclavicular region (left panel: coronal view; right panel: transverse view) of one 73
subject during monthly temperature acclimatization. BAT (Hounsfield units: -300 to -10 74
and SUV>2) was shown in red. Panels E-G show fold changes of BAT volume [Panel E], 75
mean SUV [Panel F] and BAT activity [Panel G] relative to month 1 (24°C) across 4 76
months of acclimatization. Fold change in whole fat activity, as defined by
18
F-77
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8
fluodeoxyglucose uptake within tissue of fat density (Hounsfield units: -300 to -10), is 78
displayed in [Panel H]. Two subjects [Figure S5 and S7] had little visible BAT using a 79
SUV threshold of 2.0. However, both subjects manifested an increase in whole fat 80
activity, indicating enhancement of fat metabolism [Panel H], but the overall level did not 81
reach the SUV threshold. This is illustrated in Figure S8, showing PET-CT images with 82
lower SUV threshold (1.0), and the temperature-dependent effects on fat activity. 83
84
Figure S8 Temperature-dependent fat activity of subjects with low BAT status using 85
lower SUV threshold. Panel A and Panel B show re-analysis PET-CT images of two 86
subjects with little visible BAT using a SUV threshold of 2 (see Figure S5 and S7) using 87
a SUV threshold of 1. Adipose tissue (Hounsfield unit: -300 to -10) is shown in green in 88
coronal CT sections on the left column, which display the major supraclavicular fat depot 89
at the sterno-clavicular joint. Adipose tissue with SUV1 is shown in orange in the 90
corresponding PET images in the right column. This “low-activity” BAT increased 91
following one month of cold acclimatization (19°C), decreased to nearly baseline level 92
after thermoneutral month (24°C), and was completely abolished at the end of 1 month 93
warm exposure in the final month (27°C) in both subjects. These results are concordant 94
with overall changes in fat metabolic activity during acclimatization, shown in Panel H of 95
Figure S5 and S7. 96
97
Figure S9 Summary of individual perception to cold sensation during study period 98
Participants reported subjectively their perception to cold sensation (how cold do you 99
feel) at the end of each month after 24 hour exposure to 19°C on a visual analogue scale 100
Page 47 of 59 Diabetes
9
from 1 (not cold at all) to 10 (extremely cold). Perception to cold did not change during 101
monthly acclimation. 102
103
Figure S10 Summary of appetite visual analogue scale questionnaires Panels A-C 104
(pre-meal) and D-F (post-meal) show mean scores of the three displayed questions 105
obtained from bi-weekly questionnaires. Panels G-I show AUC scores to the three 106
questions obtained during monthly ad libitum meal test. No significant trends were 107
observed in biweekly questionnaires. However, during ad libitum meal test, there was a 108
strong trend for subjects to report increased hunger [Panel G] and decreased satiety 109
[Panel H] following cold acclimatization (month 2), with scores returning to baseline 110
following sequential re-warming during months 3 and 4. Scores reflecting changes in 111
desire to eat were concordant to hunger and satiety scores, reached significance across 4 112
month acclimatization [Panel I]. *p<0.05, compared to month 1 and
#
p<0.05, compared 113
to month 2. 114
115
116
117
118
119
120
121
122
123
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Figure S1 124
125
126
127
128
129
130
131
132
133
134
135
136
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Figure S2 137
138
139
140
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Figure S3 141
142
143
144
145
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Figure S4 146
147
148
149
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Figure S5 150
151
152
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Figure S6 153
154
155
156
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Figure S7 157
158
159
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Figure S8 160
161
162
163
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Figure S9 164
165
166
167
168
169
170
171
172
173
174
175
176
177
178
179
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Figure S10 180
181
182
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References: 183
184
1. Celi FS, Brychta RJ, Linderman JD, et al. Minimal changes in environmental 185
temperature result in a significant increase in energy expenditure and changes in 186
the hormonal homeostasis in healthy adults. Eur J Endocrinol 2010;163:863-72. 187
2. Brychta RJ, Rothney MP, Skarulis MC, Chen KY. Optimizing energy 188
expenditure detection in human metabolic chambers. Conf Proc IEEE Eng Med Biol 189
Soc 2009;2009:6864-8. 190
3. de Jonge L, Nguyen T, Smith SR, Zachwieja JJ, Roy HJ, Bray GA. Prediction of 191
energy expenditure in a whole body indirect calorimeter at both low and high levels 192
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accelerometer. J Appl Physiol 1997;83:2112-22. 197
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Supplementary resource (1)

... SVC are a dynamic and complex assortment of resident immune cells, vascular cells, mesenchymal stromal cells (MSCs), and preadipocytes that change with WAT development and remodeling (Eto et al, 2009). Platelet-derived growth factor receptors (PDGFRs) α and β mark the adipose progenitor cells in the stromalvascular niche (Berry & Rodeheffer, 2013;Lee et al, 2014;Vishvanath et al, 2016). Indeed, multiple studies have reported that the PDGFRα lineage generates most of the adipocytes in response to adipogenic stimuli (Berry & Rodeheffer, 2013;Lee et al, 2014). ...
... Platelet-derived growth factor receptors (PDGFRs) α and β mark the adipose progenitor cells in the stromalvascular niche (Berry & Rodeheffer, 2013;Lee et al, 2014;Vishvanath et al, 2016). Indeed, multiple studies have reported that the PDGFRα lineage generates most of the adipocytes in response to adipogenic stimuli (Berry & Rodeheffer, 2013;Lee et al, 2014). In contrast, studies of the PDGFRβ lineage reached different conclusions (Vishvanath et al, 2016). ...
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Adipose tissue fibrosis is regulated by the chronic and progressive metabolic imbalance caused by differences in caloric intake and energy expenditure. By exploring the cellular heterogeneity within fibrotic adipose tissue, we demonstrate that early adipocyte progenitor cells expressing both platelet-derived growth factor receptor (PDGFR) α and β are the major contributors to extracellular matrix deposition. We show that the fibrotic program is promoted by senescent macrophages. These macrophages were enriched in the fibrotic stroma and exhibit a distinct expression profile. Furthermore, we demonstrate that these cells display a blunted phagocytotic capacity and acquire a senescence-associated secretory phenotype. Finally, we determined that osteopontin, which was expressed by senescent macrophages in the fibrotic environment promoted progenitor cell proliferation, fibrotic gene expression, and inhibited adipogenesis. Our work reveals that obesity promotes macrophage senescence and provides a conceptual framework for the discovery of rational therapeutic targets for metabolic and inflammatory disease associated with obesity.
... Having a higher heat intolerance and a lower cold intolerance might indicate a lower location of the thermal comfort zone or the thermoneutral zone. Recent studies have reported that cold exposure below the lower margin of the thermoneutral zone (lower critical temperature, LCT) increases thermogenesis and affects insulin sensitivity [8,50]. In those in whom LCT is lower than that in others, a relatively colder environment is needed to elicit CIT, and a relatively mild cold exposure (above their LCT and below others' LCT) might not cause thermogenesis, whereas it causes thermogenesis in others. ...
... In the recent years, CIT has gained considerable interest and has been proposed as a potential treatment for metabolic disorders, as it leads to an increase in energy expenditure, oxidation of glucose and triglycerides as substrates, and insulin sensitivity enhancement [50,60,61]; Although lifestyle programs involving exercise or diet are effective for metabolic disorders, long-term adherence is often not realized, justifying the need for other approaches, such as environmental adjustment (i.e., cold exposure) [8]. However, the duration, timing, and temperatures for CIT that are most effective to induce an increase in thermogenesis and are thus treat metabolic diseases have not yet been determined [62]. ...
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A possible association between metabolic disorders and ambient temperature has been suggested, and cold exposure as a way of increasing energy expenditure has gained considerable interest for preventative/therapeutic measures toward metabolic disorders. Although thermal sensitivity, which has recently been studied in regard to its utility as a risk assessment/patient stratification for various diseases, might influence physiological responses to ambient temperature on an individual basis, more studies are needed. We aimed to investigate the association between self-identified thermal intolerance/sensation and metabolic syndrome (MetS) to verify the working hypothesis that individuals with altered thermal sensitivity may have a predisposition to MetS. We fitted generalized additive models for thermal intolerance/sensation using body mass index (BMI) and waist-hip ratio in women, and identified those with higher/lower thermal intolerance/sensation than those predicted by the models. Higher heat intolerance, higher heat sensation, and lower cold intolerance were associated with a higher prevalence of MetS. The risk of having MetS was increased in those who had two or three associated conditions compared with those with none of these conditions. In an analysis for MetS components, significant associations of thermal sensitivity were present with high glucose, triglyceride, and blood pressure levels. Overall, higher heat intolerance/sensation and lower cold intolerance were associated with increased prevalence of MetS even at a similar level of obesity. Our study indicates that evaluation of thermal sensitivity may help identify individuals at high risk for MetS, and lead to more advanced patient stratification and personalized treatment strategies for MetS, including cold-induced thermogenesis. Supplementary information: The online version contains supplementary material available at 10.1007/s13167-022-00273-6.
... Acute cold exposure increases the uptake and oxidation of fatty acids (FA) and glucose by BAT, associated with an increase in energy expenditure in humans (EE) (10,11). In addition, cold acclimation improves peripheral insulin sensitivity in humans (12,13), likely explained by a combination of increased thermogenesis in skeletal muscle (12) and BAT (13,14). Compared to rodents, humans possess limited amounts of BAT (approx. ...
... Acute cold exposure increases the uptake and oxidation of fatty acids (FA) and glucose by BAT, associated with an increase in energy expenditure in humans (EE) (10,11). In addition, cold acclimation improves peripheral insulin sensitivity in humans (12,13), likely explained by a combination of increased thermogenesis in skeletal muscle (12) and BAT (13,14). Compared to rodents, humans possess limited amounts of BAT (approx. ...
Article
Context Cold exposure mobilizes lipids to feed thermogenic processes in organs, including brown adipose tissue (BAT). In rodents, BAT metabolic activity exhibits a diurnal rhythm, which is highest at the start of the wakeful period. Objective To investigate whether cold-induced thermogenesis displays diurnal variation in humans, and differs between males and females. Design Randomized crossover study. Participants Twenty-four young and lean males (n=12) and females (n=12). Intervention 2.5-hour personalized cooling using water-perfused mattresses in the morning (7:45 AM) and evening (7:45 PM), with one day in between. Main outcome measures Energy expenditure (EE) and supraclavicular skin temperature in response to cold exposure. Results In males, cold-induced EE was higher in the morning than in the evening (+54±10% vs. +30±7%, P=0.05). By contrast, cold-induced EE did not differ between the morning and the evening in females (+37±9% vs. +30±10%, P=0.42). Additionally, only in males, supraclavicular skin temperature upon cold increased more in the morning than in the evening (+0.2±0.1°C vs. -0.2±0.2°C, P=0.05). In males, circulating free fatty acid (FFA) levels were increased after cold in the morning, but not in the evening (+90±18% vs. +9±8%, P<0.001). In females, circulating FFA (+94±21% vs. +20±5%, P=0.006), but also triglycerides (+42±5% vs. +29±4%, P=0.01) and cholesterol levels (+17±2% vs. 11±2%, P=0.05) were more increased after cold exposure in the morning, than in the evening. Conclusions Cold-induced thermogenesis is higher in the morning than in the evening in males, however, lipid metabolism is more modulated in the morning than in the evening in females.
... Exposure to cold stimulates energy use through activation of brown adipose tissue, which contributes to thermo genesis via uncoupling mechanisms 67 . Elevated brown adipose tissue activity has been linked with improved glycaemic control and insulin sensitivity in both healthy individuals and patients with type 2 diabetes [68][69][70] . At the population level, high mean annual temperature was associated with elevated fasting plasma glucose levels, insulin resistance, and increased incidence and prevalence of diabetes [71][72][73] (Supplementary Table 1). ...
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Climate change is the greatest existential challenge to planetary and human health and is dictated by a shift in the Earth’s weather and air conditions owing to anthropogenic activity. Climate change has resulted not only in extreme temperatures, but also in an increase in the frequency of droughts, wildfires, dust storms, coastal flooding, storm surges and hurricanes, as well as multiple compound and cascading events. The interactions between climate change and health outcomes are diverse and complex and include several exposure pathways that might promote the development of non-communicable diseases such as cardiovascular disease. A collaborative approach is needed to solve this climate crisis, whereby medical professionals, scientific researchers, public health officials and policymakers should work together to mitigate and limit the consequences of global warming. In this Review, we aim to provide an overview of the consequences of climate change on cardiovascular health, which result from direct exposure pathways, such as shifts in ambient temperature, air pollution, forest fires, desert (dust and sand) storms and extreme weather events. We also describe the populations that are most susceptible to the health effects caused by climate change and propose potential mitigation strategies, with an emphasis on collaboration at the scientific, governmental and policy levels. The relationship between climate change and health outcomes is complex. In this Review, Rajagopalan and colleagues describe the environmental exposures associated with climate change and provide an overview of the consequences of climate change, including air pollution and extreme temperatures, on cardiovascular health and disease.
... The lower blood glucose levels in patients with AIS in the highlands may be related to the cold climate increasing the human metabolic rate and the expression of hypoxia-inducible transcription factor (HIF). Lee's study found that ten h of daily cold exposure at 19 • C room temperature increased human insulin sensitivity, glucose uptake, and metabolic rate after 1 month (32). Genome-wide scans of highlanders exposed to chronic hypoxia showed that the HIF signaling pathway is highly expressed (33): HIF-1α acts in skeletal muscle to increase glucose utilization and glycolysis; HIF-2α acts mainly in the liver to inhibit hepatic gluconeogenesis, ultimately resulting in significantly lower fasting glucose levels in chronically exposed highlanders compared to the plains (34). ...
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Background and Purpose Acute ischemic stroke has a high incidence in the plateau of China. It has unique characteristics compared to the plains, and the specific relationship with altitude has not yet been appreciated. This study aimed to investigate the specificity of the plateau's anterior circulation acute ischemic stroke in China. Methods To retrospectively collect clinical data of patients with first-episode acute ischemic stroke in the anterior circulation in Tianjin and Xining city. The differences in clinical presentation, laboratory, and imaging examinations were compared. Results Patients at high altitudes showed a significant trend toward lower age (61.0 ± 10.2 vs. 64.8 ± 8.1, P = 0.010) and had a history of dyslipidemia, higher levels of inflammatory markers, erythrocytosis, and alcohol abuse. The main manifestations were higher diastolic blood pressure (85.5 ± 14.0 mmHg vs. 76.8 ± 11.6 mmHg, P < 0.001), triglycerides [2.0 (1.8) mmol/L vs. 1.3 (0.9) mmol/L, P < 0.001], CRP [4.7 (4.4) mg/L vs. 2.1 (1.9) mg/L, P < 0.001], homocysteine levels [14.5 (11.7) μmol/L vs. 11.2 (5.2) μmol/L, P < 0.001]; larger infarct volume [3.5 (4.8) cm ³ vs. 9.0 (6.9) cm ³ , P < 0.001] and worse prognosis. Patients at high altitudes had higher atherosclerotic indexes in cIMT and plaque than those in plains. Conclusions The natural habituation and genetic adaptation of people to the particular geo-climatic environment of the plateau have resulted in significant differences in disease characteristics. Patients with the anterior circulation acute ischemic stroke in the plateau show more unfavorable clinical manifestations and prognosis. This study provides a preliminary interpretation of the effects of altitude and suggests developing preventive and therapeutic protocol measures that are more appropriate for the plateau of China.
... The host provides a habitat for specific intestinal microbial communities, which in turn provide nutrition and energy for the host and promote the acquisition of food energy and the storage of white fat [2][3][4] . Changes in ambient temperature are among the strongest physiological stimuli that increase the formation and activity of thermogenic fat 5,6 . In mammals, nonshivering thermogenesis (NST) generates metabolic heat to maintain body temperature in response to cold 7 . ...
Article
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The gut microbiota plays a key role in host metabolic thermogenesis by activating UCP1 and increasing the browning process of white adipose tissue (WAT), especially in cold environments. However, the crosstalk between the gut microbiota and the host, which lacks functional UCP1, making them susceptible to cold stress, has rarely been illustrated. We used male piglets as a model to evaluate the host response to cold stress via the gut microbiota (four groups: room temperature group, n = 5; cold stress group, n = 5; cold stress group with antibiotics, n = 5; room temperature group with antibiotics, n = 3). We found that host thermogenesis and insulin resistance increased the levels of serum metabolites such as glycocholic acid (GCA) and glycochenodeoxycholate acid (GCDCA) and altered the compositions and functions of the cecal microbiota under cold stress. The gut microbiota was characterized by increased levels of Ruminococcaceae, Prevotellaceae, and Muribaculaceae under cold stress. We found that piglets subjected to cold stress had increased expression of genes related to bile acid and short-chain fatty acid (SCFA) metabolism in their liver and fat lipolysis genes in their fat. In addition, the fat lipolysis genes CLPS, PNLIPRP1, CPT1B, and UCP3 were significantly increased in the fat of piglets under cold stress. However, the use of antibiotics showed a weakened or strengthened cold tolerance phenotype, indicating that the gut microbiota plays important role in host thermogenesis. Our results demonstrate that the gut microbiota-blood-liver and fat axis may regulate thermogenesis during cold acclimation in piglets.
... Most importantly, the BAT-mediated improvements in metabolic health are more prominent in overweight and obese individuals [22], indicating that BAT can disengage obesity from disease. Thus, while the potential for BAT in treating human obesity can be debated, strong experimental evidence supports the idea that BAT can offset the adverse effects of obesity [22][23][24][25]. ...
Chapter
Thermogenic adipose tissue plays a vital function in regulating whole-body energy expenditure and nutrient homeostasis due to its capacity to dissipate chemical energy as heat, in a process called non-shivering thermogenesis. A reduction of creatine levels in adipocytes impairs thermogenic capacity and promotes diet-induced obesityKazak et al, Cell 163, 643–55, 2015; Kazak et al, Cell Metab 26, 660–671.e3, 2017; Kazak et al, Nat Metab 1, 360–370, 2019). Mechanistically, thermogenic respiration can be promoted by the liberation of an excess quantity of ADP that is dependent on addition of creatine. A model of a two-enzyme system, which we term the Futile Creatine Cycle, has been posited to support this thermogenic action of creatine. Futile creatine cycling can be monitored in purified mitochondrial preparations wherein creatine-dependent liberation of ADP is monitored through the measurement of oxygen consumption under ADP-limiting conditions. The current model proposes that, in thermogenic fat cells, mitochondria-targeted creatine kinase B (CKB) uses mitochondrial-derived ATP to phosphorylate creatine (Rahbani JF, Nature 590, 480–485, 2021). The creatine kinase reaction generates phosphocreatine and ADP, and ADP stimulates respiration. Next, a pool of mitochondrial phosphocreatine is directly hydrolyzed by a phosphatase, to regenerate creatine. The liberated creatine can then engage mitochondrial CKB to trigger another round of this cycle to support ADP-dependent respiration. In this model, the coordinated action of creatine phosphorylation and phosphocreatine hydrolysis triggers a futile cycle that produces a molar excess of mitochondrial ADP to promote thermogenic respiration (Rahbani JF, Nature 590, 480–485, 2021; Kazak and Cohen, Nat Rev Endocrinol 16, 421–436, 2020). Here, we provide a detailed method to perform respiratory measurements on isolated mitochondria and calculate the stoichiometry of creatine-dependent ADP liberation. This method provides a direct measure of the futile creatine cycle.
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The discovery of functional brown adipose tissue (BAT) in adult humans and the possibility to recruit beige cells with high thermogenic potential within white adipose tissue (WAT) depots opened the field for new strategies to combat obesity and its associated comorbidities. Exercise training as well as cold exposure and dietary components are associated with the enhanced accumulation of metabolically-active beige adipocytes and BAT activation. Both activated beige and brown adipocytes increase their metabolic rate by utilizing lipids to generate heat via non-shivering thermogenesis, which is dependent on uncoupling protein 1 (UCP1) in the inner mitochondrial membrane. Non-shivering thermogenesis elevates energy expenditure and promotes a negative energy balance, which may ameliorate metabolic complications of obesity and Type 2 Diabetes Mellitus (T2DM) such as insulin resistance (IR) in skeletal muscle and adipose tissue. Despite the recent advances in pharmacological approaches to reduce obesity and IR by inducing non-shivering thermogenesis in BAT and WAT, the administered pharmacological compounds are often associated with unwanted side effects. Therefore, lifestyle interventions such as exercise, cold exposure, and/or specified dietary regimens present promising anchor points for future disease prevention and treatment of obesity and T2DM. The exact mechanisms where exercise, cold exposure, dietary interventions, and pharmacological treatments converge or rather diverge in their specific impact on BAT activation or WAT browning are difficult to determine. In the past, many reviews have demonstrated the mechanistic principles of exercise- and/or cold-induced BAT activation and WAT browning. In this review, we aim to summarize not only the current state of knowledge on the various mechanistic principles of diverse external stimuli on BAT activation and WAT browning, but also present their translational potential in future clinical applications.
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Brown adipose tissue (BAT) is a metabolically active organ that contributes to the thermogenic response to cold exposure. In addition, other thermogenic cells termed beige adipocytes are generated in white adipose tissue (WAT) by cold exposure. While activation of brown/beige adipose tissue is associated with mobilization of both glucose and lipids, few studies have focused on the role of glycolytic enzymes in regulating adipose tissue function. We generated mouse models with specific deletion of the glycolytic enzyme phosphoglycerate mutase 1 (PGAM1) from adipose tissue. Deletion of Pgam1 from both BAT and WAT promoted whitening of BAT with beiging of visceral WAT, whereas deletion of Pgam1 from BAT alone led to whitening of BAT without beiging of WAT. Our results demonstrate a potential role of glycolytic enzymes in beiging of visceral WAT and suggest that PGAM1 would be a novel therapeutic target in obesity and diabetes.
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Obesity is a chronic condition of multifactorial etiology characterized by excessive body fat due to a calorie intake higher than energy expenditure. Given the intrinsic limitations of surgical interventions and the difficulties associated with lifestyle changes, pharmacological manipulation is currently one of the main therapies for metabolic diseases. Approaches aiming to promote energy expenditure through induction of thermogenesis have been explored and, in this context, brown adipose tissue (BAT) activation and browning have been shown to be promising strategies. Although such processes are physiologically stimulated by the sympathetic nervous system, not all situations that are known to increase adrenergic signaling promote a concomitant increase in BAT activation or browning in humans. Thus, a better understanding of factors involved in the thermogenesis attributed to these tissues is needed to enable the development of future therapies against obesity. Herein we carry out a critical review of original articles in humans under conditions previously known to trigger adrenergic responses—namely, cold, catecholamine-secreting tumor (pheochromocytoma and paraganglioma), burn injury, and adrenergic agonists—and discuss which of them are associated with increased BAT activation and browning. BAT is clearly stimulated in individuals exposed to cold or treated with high doses of the β3-adrenergic agonist mirabegron, whereas browning is certainly induced in patients after burn injury or with pheochromocytoma, as well as in individuals treated with β3-adrenergic agonist mirabegron for at least 10 weeks. Given the potential effect of increasing energy expenditure, adrenergic stimuli are promising strategies in the treatment of metabolic diseases.
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The simultaneous assessment of insulin action, secretion, and hepatic extraction is key to understanding postprandial glucose metabolism in nondiabetic and diabetic humans. We review the oral minimal method (i.e., models that allow the estimation of insulin sensitivity, β-cell responsivity, and hepatic insulin extraction from a mixed-meal or an oral glucose tolerance test). Both of these oral tests are more physiologic and simpler to administer than those based on an intravenous test (e.g., a glucose clamp or an intravenous glucose tolerance test). The focus of this review is on indices provided by physiological-based models and their validation against the glucose clamp technique. We discuss first the oral minimal model method rationale, data, and protocols. Then we present the three minimal models and the indices they provide. The disposition index paradigm, a widely used β-cell function metric, is revisited in the context of individual versus population modeling. Adding a glucose tracer to the oral dose significantly enhances the assessment of insulin action by segregating insulin sensitivity into its glucose disposal and hepatic components. The oral minimal model method, by quantitatively portraying the complex relationships between the major players of glucose metabolism, is able to provide novel insights regarding the regulation of postprandial metabolism.
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Excess lipid storage in adipose tissue results in the development of obesity and other metabolic disorders including diabetes, fatty liver and cardiovascular diseases. The lipid droplet (LD) is an important subcellular organelle responsible for lipid storage. We previously observed that Fsp27, a member of the CIDE family proteins, is localized to LD-contact sites and promotes atypical LD fusion and growth. Cidea, a close homolog of Fsp27, is expressed at high levels in brown adipose tissue. However, the exact role of Cidea in promoting LD fusion and lipid storage in adipose tissue remains unknown. Here, we expressed Cidea in Fsp27-knockdown adipocytes and observed that Cidea has similar activity to Fsp27 in promoting lipid storage and LD fusion and growth. Next, we generated Cidea and Fsp27 double-deficient mice and observed that these animals had drastically reduced adipose tissue mass and a strong lean phenotype. In addition, Cidea/Fsp27 double-deficient mice had improved insulin sensitivity and were intolerant to cold. Furthermore, we observed that the brown and white adipose tissues of Cidea/Fsp27 double-deficient mice had significantly reduced lipid storage and contained smaller LDs compared to those of Cidea or Fsp27 single deficient mice. Overall, these data reveal an important role of Cidea in controlling lipid droplet fusion, lipid storage in brown and white adipose tissue, and the development of obesity.
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In recent years, it has been shown that humans have active brown adipose tissue (BAT) depots, raising the question of whether activation and recruitment of BAT can be a target to counterbalance the current obesity pandemic. Here, we show that a 10-day cold acclimation protocol in humans increases BAT activity in parallel with an increase in nonshivering thermogenesis (NST). No sex differences in BAT presence and activity were found either before or after cold acclimation. Respiration measurements in permeabilized fibers and isolated mitochondria revealed no significant contribution of skeletal muscle mitochondrial uncoupling to the increased NST. Based on cell-specific markers and on uncoupling protein-1 (characteristic of both BAT and beige/brite cells), this study did not show "browning" of abdominal subcutaneous white adipose tissue upon cold acclimation. The observed physiological acclimation is in line with the subjective changes in temperature sensation; upon cold acclimation, the subjects judged the environment warmer, felt more comfortable in the cold, and reported less shivering. The combined results suggest that a variable indoor environment with frequent cold exposures might be an acceptable and economic manner to increase energy expenditure and may contribute to counteracting the current obesity epidemic.
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Background The American Diabetes Association (ADA) recommend that fasting glucose alone with the oral glucose tolerance test should be used to diagnose diabetes mellitus. We assessed mortality associated with the ADA fasting-glucose criteria compared with the WHO 2 h post-challenge glucose criteria. Methods We assessed baseline data on glucose concentrations at fasting and 2 h after the 75 g oral glucose tolerance test from 13 prospective European cohort studies, which included 18 048 men and 7316 women aged 30 years or older. Mean follow-up was 7–3 years. We assessed the risk of death according to the different diagnostic glucose categories. Findings Compared with men who had normal fasting glucose (<6.1 mmol/L), men with newly diagnosed diabetes mellitus by the ADA fasting criteria (≤7.0 mmol/L) had a hazard ratio for death of 1.81 (95% CI1.49–2.20); for women the hazaid ratio was 1.79 (1.18–2.69). For impaired fasting glucose (6.1–6.9 mmol/L), the hazard ratios were 1.21 (1.05–1.41) and 1.08 (0.70–1.66). Fbrthe WHO criteria (≤11.1 mmol/L), the ratios for newly diagnosed diabetes were 2.02 (1.66–2.46) in men and 2.77 (1.96–3.92) in women, and for impaired glucose tolerance (7.8–11.1 mmol/L) were 1.51 (1.32–1.72) and 1.60 (1.22–2.10). Within each fasting-glucose classification, mortality increased with increasing 2 h glucose. However, for 2 h glucose classifications of impaired glucose tolerance, and diabetes, there was no trend for increasing fasting glucose concentrations. Interpretation Fasting-glucose concentrations alone do not identify individuals at increased risk of death associated with hyperglycaemia. The oral glucose tolerance test provides additional prognostic information and enables detection of individuals with impaired glucose tolerance, who have the greatest attributable risk of death
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Rediscovery of cold-activated brown adipose tissue (BAT) in humans has boosted research interest in identifying BAT activators for metabolic benefits. Of particular interest are cytokines capable of fat browning. Irisin, derived from FNDC5, is an exercise-induced myokine that drives brown-fat-like thermogenesis in murine white fat. Here we explored whether cold exposure is an afferent signal for irisin secretion in humans and compared it with FGF21, a brown adipokine in rodents. Cold exposure increased circulating irisin and FGF21. We found an induction of irisin secretion proportional to shivering intensity, in magnitude similar to exercise-stimulated secretion. FNDC5 and/or FGF21 treatment upregulated human adipocyte brown fat gene/protein expression and thermogenesis in a depot-specific manner. These results suggest exercise-induced irisin secretion could have evolved from shivering-related muscle contraction, serving to augment brown fat thermogenesis in concert with FGF21. Irisin-mediated muscle-adipose crosstalk may represent a thermogenic, cold-activated endocrine axis that is exploitable in obesity therapeutics development.
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There has been an upsurge of interest in the adipocyte coincident with the onset of the obesity epidemic and the realization that adipose tissue plays a major role in the regulation of metabolic function. The past few years, in particular, have seen significant changes in the way that we classify adipocytes and how we view adipose development and differentiation. We have new perspective on the roles played by adipocytes in a variety of homeostatic processes and on the mechanisms used by adipocytes to communicate with other tissues. Finally, there has been significant progress in understanding how these relationships are altered during metabolic disease and how they might be manipulated to restore metabolic health.
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Brown adipose tissue (BAT) is the site of sympathetically activated adaptive thermognenesis during cold exposure and after hyperphagia, thereby controlling whole-body energy expenditure (EE) and body fat. Radionuclide imaging studies have demonstrated that adult humans have metabolically active BAT composed of mainly beige/brite adipocytes, recently identified brown-like adipocytes. The inverse relationship between the BAT activity and body fatness suggests that BAT is, because of its energy dissipating activity, protective against body fat accumulation in humans as it is in small rodents. In fact, either repeated cold exposure or daily ingestion of some food ingredients acting on transient receptor potential channels recruits BAT in parallel with increased EE and decreased body fat. In addition to the sympathetic nervous system, several endocrine factors are also shown to recruit BAT. Thus, BAT is a promising therapeutic target for combating human obesity and related metabolic disorders.
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Background: Brown adipose tissue (BAT) is involved in the regulation of whole-body energy expenditure and adiposity. Some clinical studies have reported an association between BAT and blood glucose in humans. Objective: To examine the impact of BAT on glucose metabolism, independent of that of body fatness, age and sex in healthy adult humans. Methods: Two hundred and sixty healthy volunteers (184 males and 76 females, 20-72 years old) underwent fluorodeoxyglucose-positron emission tomography and computed tomography after 2 h of cold exposure to assess maximal BAT activity. Blood parameters including glucose, HbA1c and low-density lipoprotein (LDL)/high-density lipoprotein-cholesterol were measured by conventional methods, and body fatness was estimated from body mass index (BMI), body fat mass and abdominal fat area. The impact of BAT on body fatness and blood parameters was determined by logistic regression with the use of univariate and multivariate models. Results: Cold-activated BAT was detected in 125 (48%) out of 260 subjects. When compared with subjects without detectable BAT, those with detectable BAT were younger and showed lower adiposity-related parameters such as the BMI, body fat mass and abdominal fat area. Although blood parameters were within the normal range in the two subject groups, HbA1c, total cholesterol and LDL-cholesterol were significantly lower in the BAT-positive group. Blood glucose also tended to be lower in the BAT-positive group. Logistic regression demonstrated that BAT, in addition to age and sex, was independently associated with BMI, body fat mass, and abdominal visceral and subcutaneous fat areas. For blood parameters, multivariate analysis after adjustment for age, sex and body fatness revealed that BAT was a significantly independent determinant of glucose and HbA1c. Conclusion: BAT, independent of age, sex and body fatness, has a significant impact on glucose metabolism in adult healthy humans.