Characteristics of all subjects before and after 12-week CRD intervention
The WC, SBP, DBP, BFC, BW, BMR, HOMA-IR, TG and ALT,in all subjects significantly decreased after CRD intervention, as compared to those at baseline (Table 1).
Table 1. Characteristics of all subjects before and after 12-week CRD intervention.
Variates
|
Before CRD intervention
|
After CRD intervention
|
P
|
n
|
16
|
16
|
-
|
BMI (kg/m2)
|
30.57±2.19
|
30.52±4.45
|
0.136
|
WC (cm)
|
101.17±7.19
|
97.37±9.06
|
0.002
|
BW (kg)
|
86.94±10.85
|
81.54±12.02
|
0.002
|
SBP (mmHg)
|
125.56±9.93
|
119.31±9.74
|
0.001
|
DBP (mmHg)
|
89.31±8.81
|
82.19±5.44
|
<0.001
|
BFC (kg)
|
28.24±4.50
|
25.34±4.94
|
0.020
|
BFP (%)
|
32.55±3.93
|
30.84±4.02
|
0.085
|
BMR (kcal)
|
1489.56±187.75
|
1451.63±187.75
|
<0.001
|
VFA (cm2)
|
140.00±30.73
|
123.56±32.64
|
0.124
|
HbA1c(%)
|
5.46±.38
|
5.26±.45
|
0.168
|
HOMA-IR
|
1.54±0.39
|
1.35±0.34
|
0.017
|
TC (mmol/l)
|
4.86±0.96
|
4.38±0.87
|
0.068
|
TG (mmol/l)
|
1.62±1.19
|
1.07±0.77
|
0.009
|
ALT (U/l)
|
35.63±26.52
|
21.13±9.15
|
0.028
|
AST (U/l)
|
25.19±11.47
|
19.06±4.88
|
0.050
|
ALP (U/l)
|
68.75±26.01
|
82.88±26.03
|
0.144
|
BUN (mmol/l)
|
5.57±1.03
|
4.85±1.27
|
0.092
|
Cr (μmol/l)
|
70.53±19.31
|
67.59±13.60
|
0.401
|
eGFR (ml/min)
|
112.64±17.02
|
117.03±10.1
|
0.231
|
Notes: All variables are expressed as n (%) for qualitative data or mean ± standard deviation for quantitative data.
BMI, Body mass index; WC, Waist circumference; BW, Body weight; SBP, Systolic blood pressure; DBP, Diastolic blood pressure; BFC, Body Fat Content; BFP, Body Fat Percentage; BMR, Basal metabolic rate; VFA, Visceral Fat Area; HbA1c, Hemoglobin A1C; HOMA-IR, Homeostasis model of assessment for insulin resistance index; TC, Total cholesterol; TG, Triglycerides; ALT, Alanine aminotransferase; AST, Aspartate aminotransferase; ALP, Alkaline phosphatase; BUN, Blood urea nitrogen; Cr, creatinine; eGFR, Estimated glomerular filtration rate.
Characteristics of subjects in two groups before and after CRD intervention
At baseline, there were no marked differences in the age, gender, BMI, WC, BW, SBP, DBP, BFC, BFP, BMR, VFA, HbA1c, HOMA-RI, HbA1c, TC, TG, ALT, AST, ALP, BUN and eGFR between SD group and NSD group (Table 2). After CRD intervention, the BW, WC, BMR, SBP and DBP decreased significantly in both groups. However, there were slight declines in the TG in the SD group and TC,HOMA-RI in the NSD group (Table 2).
Table 2. Characteristics of subjects in two groups before and after intervention
Variates
|
SD
|
Pa
|
NSD
|
Pb
|
P0
|
Before CRD intervention
|
After CRD intervention
|
Before CRD intervention
|
After CRD intervention
|
n
|
8
|
-
|
-
|
8
|
-
|
-
|
-
|
Gender (Male, %)
|
7 (87.5%)
|
-
|
-
|
5 (62.5%)
|
-
|
-
|
-
|
Age (years)
|
38.13±2.44
|
-
|
-
|
34.00±3.78
|
-
|
-
|
0.160
|
BMI (kg/m2)
|
29.85±1.71
|
29.33±1.82
|
0.148
|
31.29±2.49
|
29.30±6.25
|
0.247
|
0.199
|
WC (cm)
|
100.73±6.14
|
98.60±6.76
|
0.008
|
101.61±8.52
|
96.14±11.26
|
0.017
|
0.815
|
BW (kg)
|
85.88±8.76
|
83.85±9.30
|
0.023
|
88.00±13.16
|
79.23±14.52
|
0.006
|
0.709
|
|
SBP (mmHg)
|
128.38±7.27
|
120.63±9.44
|
0.019
|
122.75±11.85
|
118.00±10.50
|
0.012
|
0. 272
|
DBP (mmHg)
|
91.38±9.41
|
82.88±5.84
|
0.005
|
87.25±8.24
|
81.50±5.32
|
0.008
|
0. 367
|
BFC (kg)
|
27.11±3.28
|
26.56±2.78
|
0.447
|
29.36±5.44
|
24.11±6.41
|
0.023
|
0.334
|
BFP (%)
|
31.64±2.95
|
31.33±2.25
|
0.724
|
33.46±4.74
|
30.36±5.38
|
0.085
|
0.371
|
BMR (kcal)
|
1501.75±149.36
|
1476.25±146.63
|
0.009
|
1477.38±229.98
|
1427.00±228.02
|
0.001
|
0.805
|
VFA (cm2)
|
135.00±19.31
|
138.25±23.34
|
0.544
|
145.00±39.92
|
108.88±35.29
|
0.075
|
0.534
|
HbA1c (%)
|
5.39±0.36
|
5.24±0.58
|
0.540
|
5.53±0.41
|
5.29±0.31
|
0.168
|
0.490
|
HOMA-IR
|
2.68±1.19
|
2.15±0.84
|
0.308
|
3.44±1.83
|
1.91±1.78
|
0.033
|
0.344
|
TC (mmol/l)
|
4.56±0.66
|
4.62±0.77
|
0.645
|
4.91±1.18
|
4.16±0. 95
|
0.024
|
0.457
|
TG (mmol/l)
|
1.71±1.33
|
1.20±1.01
|
0.040
|
1.50±1.11
|
0.94±0.45
|
0.115
|
0.742
|
ALT (U/l)
|
26.75±8.46
|
19.75±7.32
|
0.055
|
44.50±35.43
|
22.50±11.02
|
0.092
|
0.206
|
AST (U/l)
|
23.38±6.80
|
19.38±2.67
|
0.175
|
27.00±15.10
|
18.75±6.61
|
0.158
|
0.546
|
ALP (U/l)
|
60.75±20.53
|
72.88±27.2
|
0.418
|
76.75±29.68
|
92.87±21.94
|
0.242
|
0.230
|
BUN (mmol/l)
|
5.28±0.94
|
4.77±1.05
|
0.457
|
5.85±1.09
|
4.93±1.52
|
0.107
|
0.277
|
Cr (μmol/l)
|
73.12±18.67
|
72.11±11.95
|
0.855
|
67.94±20.86
|
63.08±14.37
|
0.309
|
0.609
|
eGFR (ml/min)
|
110.97±16.36
|
113.42±10.61
|
0.654
|
114.32±18.62
|
120.65±8.79
|
0.243
|
0.708
|
Notes: All variables are expressed as n (%) for qualitative data or mean ± standard deviation for quantitative data. Pa, the difference of the change of each parameter before and after intervention in the SD group. Pb, the difference of the change of each parameter before and after intervention in the NSD group. P0, the difference of baseline parameters between SD group and NSD group. NSD, non-sleep deprivation group. SD, sleep deprivation group. BMI, Body mass index; WCm Waist circumference; BWm Body weight; SBP, Systolic blood pressure; DBP, Diastolic blood pressure; BFC, Body Fat Content; BFP, Body Fat Percentage; BMR, Basal metabolic rate; VFA, Visceral Fat Area; HbA1c, Hemoglobin A1C; HOMA-IR, Homeostasis model of assessment for insulin resistance index; TC, Total cholesterol; TG, Triglycerides; ALT, Alanine aminotransferase; AST, Aspartate aminotransferase; ALP, Alkaline phosphatase; BUN, Blood urea nitrogen; Cr, creatinine; eGFR, Estimated glomerular filtration rate
Changes in the parameters in two groups before and after the CRD intervention
After intervention, the mean BFC, VFA, BW, BMR and TC in the NSD group decreased by 4.70 kg, 39.38 cm2, 6.74 kg, 24.88 kcal and 0.83 mmol/l, respectively, and these changes were more obvious than those in the SD group. However, there were no significant differences in the changes of BMI, BFP, WC, ALT, AST, ALP, BUN, eGFR, TG, HOMA-RI,HbA1c, SBP and DBP between NSD group and SD group (Table 3).
Table 3. Changes in the body parameters and biochemical parameters after CRD intervention in two groups.
Variates
|
SD
|
NSD
|
P
|
n
|
8
|
8
|
-
|
∆BMI (kg/m2)
|
0.53±0.91
|
1.99±4.45
|
0.378
|
∆WC (CM)
|
2.13±1.63
|
5.48±4.99
|
0.107
|
∆BW (kg)
|
2.02±1.97
|
8.76±6.36
|
0.013
|
∆SBP (mmHg)
|
7.75±7.25
|
4.75±3.99
|
0.327
|
∆DBP (mmHg)
|
8.50±6.00
|
5.75±4.40
|
0.314
|
∆BFC (kg)
|
0.55±1.93
|
5.25±5.10
|
0.029
|
∆BFP (%)
|
0.31±2.40
|
3.10±4.38
|
0.137
|
∆BMR (kcal)
|
25.50±20.04
|
50.38±23.77
|
0.040
|
∆VFA (cm2)
|
-3.25±14.41
|
36.13±48.94
|
0.047
|
∆HbA1c
|
0.15±0.66
|
0.23±0.42
|
0.179
|
∆HOMA-RI
|
0.53±1.36
|
1.53±1.64
|
0.904
|
∆TC (mmol/l)
|
-0.06±0.37
|
0.77±0.76
|
0.014
|
∆TG (mmol/l)
|
0.051±0.57
|
0.56±0.881
|
0.895
|
∆ALT (U/l)
|
7.00±8.59
|
22.00±31.85
|
0.219
|
∆AST (U/l)
|
4.00±7.50
|
8.25±14.75
|
0.480
|
∆ALP (U/l)
|
-12.13±39.87
|
-16.13±35.69
|
0.836
|
∆BUN (mmol/l)
|
0.51±1.82
|
0.92±1.41
|
0.615
|
∆Cr (μmol/l)
|
1.01±15.17
|
4.86±12.55
|
0.589
|
∆eGFR (ml/min)
|
2.45±14.78
|
6.33±14.03
|
0.599
|
Note: All variables are expressed as n (%) for qualitative data or mean ± standard deviation for quantitative data. NSD, non-sleep deprivation group. SD, sleep deprivation group. BMI, Body mass index; WC, Waist circumference; BW, Body weight; SBP, Systolic blood pressure; DBP, Diastolic blood pressure; BFC, Body Fat Content; BFP, Body Fat Percentage; BMR, Basal metabolic rate; VFA, Visceral Fat Area; HbA1c, Hemoglobin A1C; HOMA-IR, Homeostasis model of assessment for insulin resistance index; TC, Total cholesterol; TG, Triglycerides; ALT, Alanine aminotransferase; AST, Aspartate aminotransferase; ALP ,Alkaline phosphatase; BUN, Blood urea nitrogen; Cr, creatinine; eGFR, Estimated glomerular filtration rate. “∆”: difference between baseline level and 12-week level.
Findings from Doppler ultrasonography
At baseline, fatty liver disease was noted in all the subjects of both SD group and NSD group. After 12-week intervention, fatty liver disease was not observed in 87.5% and 37.5% of subjects in the NSD group and SD group, respectively.
Gut microbiota distribution
Sequencing depth of gut microbiota: the species accumulation curve of samples was analyzed by random sampling method to detect the sequencing depth. The results showed that the dilution curve tended to flatten when there were 30 sequencing samples, indicating that the depth of this sequencing met the requirements, and the additional amount of sequencing data was not necessary for studying the discovery of OTU (Figure 1).
Gut microbiota in two groups before and after CRD intervention
As shown in Fig.2A, the numbers of OTUs were similar between SD0 and SD12, NSD0 and NSD12, SD0 and NSD0, and between SD12 and NSD12 (PSD0-SD12=0.57, PNSD0-NSD12=0.08, PSD0-NSD0=0.29 and PSD12-NSD12=0.43). However, as shown in Fig.2B, the abundance in the NSD group increased gradually after intervention, while that in the SD group decreased gradually, and the abundance in the SD group was lower than in the NSD group before and after CRD intervention,without significance.
Heat map analysis of correlation between gut microbiota abundance and clinical factors before and after intervention in two groups
At the phylum level, heat maps were plotted based on the correlation coefficient between abundance of gut microbiota and clinical indicators. As shown in Figure 3, at baseline, in the NSD group, Selenomonadales was negatively related to AST, Bifidobacteriaceae and Cr. Coriobacteriaceae were negatively related to FBG. Desulfovibrionaceae was positively related to BFR. There was a negative correlation between Mollicutes and BMR.
However, there was no significant association in the SD group between abundance of gut microbiota and clinical factors.
At the end of 12 weeks after CRD intervention (Fig.4), in the NSD group, there was a significant positive correlation between Akkermansia and FBG. Bifidobacteriaceae was positively related to FBG, HbA1C and TG. Clostridiales was negatively related to BUN. There was a negative correlation between Mollicutes and BMR. However, in the SD group, Burkholderiales was positively related to FBG. Desulfovibrionaceae was positively related to AU. Actinobacteria and Actinomycetaceae were negatively related to VFA.
Mollicutes Alpha diversity in two groups before and after intervention.
A stable negative correlation between Mollicutes bacteria and BMR was noted in the NSD group before and after CRD intervention (Fig3-4). To better analyze the relations between Mollicutes and BMR, Mollicutes Alpha diversity was analyzed separately in the Figure 5. There were no significant difference in the relative abundance of Mollicutes before and after intervention in the SD group and NSD group (PSD0-SD12=0.685, PNSD0-NSD12=0.524) , However, the change of relative abundance of Mollicutes in the NSD group after intervention was significantly greater than in the SD group.