Bo′s abdominal acupuncture improves disordered metabolism in obese type 2 diabetic rats through regulating fibroblast growth factor 21 and its related adipokines

doi:10.19852/j.cnki.jtcm.20231008.002

Abstract

Abstract:

OBJECTIVE: To investigate the effect of Bo′s abdominal acupuncture (BOAA) on fibroblast growth factor 21 (FGF21) and its related adipokines in type 2 diabetes mellitus (T2DM) rats.
METHODS: This study established obese T2DM rat model by high-fat diet (HFD) with a dose of streptozotocin (STZ, 30 mg/kg). Obese T2DM rats were randomly subdivided into four groups (n = 10): negative, BOAA, conventional acupuncture (COA group) and metformin group (Met group) groups. The biochemical parameters, mRNAs, and proteins were analyzed using enzyme-lined immunoassays kits, quantitative polymerase chain reaction and Western blot.
RESULTS: Treatment with BOAA attenuated the histopathological changes in visceral fat and restored the alterations in the levels of body weight, fasting blood glucose (FBG), homeostasis model assessment for insulin resistance (HOMA-IR). BOAA treatment significantly decreased the levels of triglyceride, total cholesterol, low density lipoprotein cholesterol, leptin, and increased the serum levels of high-density lipoprotein cholesterol, fibroblast growth factor 21 (FGF21), adiponectin (ADP), peroxisome proliferator-activated receptor γ (PPAR-γ), C-peptide (C-P) in obese T2DM rats. Furthermore, BOAA treatment significantly increased the mRNA expressions of FGF21, ADP, leptin, PPAR-γ, PPAR-α and adenosine 5‘-monophosphate (AMP)-activated protein kinase (AMPK). Besides, BOAA treatment upregulated the protein expressions of fibroblast growth factor receptors3 (FGFR3), PPAR-α, extracellular signal-regulated kinase (ERK), phosphorylated ERK (p-ERK), AMPK, p-AMPK, Liver kinase B1 (LKB1), phosphorylated LKB1 (p-LKB1), acetyl-CoA carboxylase (ACC) and phosphorylated ACC (p-ACC), while downregulated the protein expressions of FGF21 and PPAR-γ in visceral fat.
CONCLUSIONS: BOAA treatment reduced FBG and body weight, and improved insulin sensitivity through regulating FGF21 signaling pathway and its related adipokine in obese T2DM rats.

Key words: diabetes mellitus, type 2, obesity, fibroblast growth factors, adipokines, insulin resistance, Bo′s abdominal acupuncture

QIN Xihui, PANG Jianli, XIONG Guan, FENG Jie. Bo′s abdominal acupuncture improves disordered metabolism in obese type 2 diabetic rats through regulating fibroblast growth factor 21 and its related adipokines[J]. Journal of Traditional Chinese Medicine, 2023, 43(6): 1200-1208.

Figures/Tables 7

Table 1 qPCR reaction condition

Step	Temperature (℃)	Time	Cycle
UDG activation	50	2 min	Hold
Dual-lock DNA polymerase	95	2 min	Hold
Denature	95	15 s	40
Anneal/extend	60	1 min	40

Table 2 Primer sequence and Tm of each gene

Gene	Sequence	Tm (℃)
FGF21	Forward Primer 5′-TCTCCTGCTGCCTGTCTTCCTG-3′	61.9
FGF21	Reverse Primer 5′-TCGGTGTCCTGGTCGTCATCTG-3′	61.7
PPAR-γ	Forward Primer 5′-CGCCAAGGTGCTCCAGAAGATG-3′	61.4
PPAR-γ	Reverse Primer 5′- AGGGTGAAGGCTCATATCTGTCTCC-3′	60.3
PPAR-α	Forward Primer 5′-GCCAAGAGAATCCACGAAGCCTAC-3′	60.3
PPAR-α	Reverse Primer 5′-TGTTGCTAGTCTTTCCTGCGAGTATG-3′	58.6
Leptin	Forward Primer 5′-GTTCCTGTGGCTTTGGTCCTATCTG-3′	59.9
Leptin	Reverse Primer 5′- TGATCCTGGTGACAATGGTCTTGATG-3′	58.8
ADP	Forward Primer 5′- ATGTATCACTCAGCATTCAGCGTAGG-3′	58.5
ADP	Reverse Primer 5′-CTGCCGTCATAATGATTCTGTTGGTTG-3′	58.2
AMPK	Forward Primer 5′- ATGATGAGGTGGTGGAGCAGAGG-3′	61.5
AMPK	Reverse Primer 5′-GGTTCTCGGCTGTGCTGGAATC-3′	60.7
β-actin	Forward Primer 5′-GGTTCTCGGCTGTGCTGGAATC-3′	60.6
β-actin	Reverse Primer 5′- AGGAAGAGGATGCGGCAGTGG-3′	62.9

Table 3 Levels of FBG, HOMA-IR and ISI

Group	n	FBG (mmol/L)	HOMA-IR	ISI
Control	10	5.00±0.15	2.38±0.19	-3.98±0.08
Negative	10	18.98±0.57^a	11.73±0.79^a	-5.58±0.07^a
Met	10	6.92±0.25^b	3.61±0.16^b	-4.40±0.05^b
BOAA	10	9.58±0.53^bc	5.28±0.42^bc	-4.70±0.12^bc
COA	10	16.40±0.73^b	9.19±0.87^b	-5.37±0.07^b

Figure 1 Effects of the body weight, blood glucose, FINS, FFA, TG, TC, LDL-C and HDL-C levels in different groups A: body weight; B: blood glucose; C: FINS; D: FAA; E: TG; F: TC; G: LDL-C; H: HDL-C. BT: before treatment; AT: after treatment. Control group: no treatment is given; negative group: treated with high fat feed; Met group: treated with metformin; COA group: treated with COA; BOAA group: treated with BOAA. BOAA: Bo′s abdominal acupuncture; COA: conventional acupuncture; FBG: fasting blood glucose; FINS: fasting insulin; FFA: free fatty acids; TC: total cholesterol; TG: total triglycerides; HDL-C: high-density lipoprotein; LDL-C: low-density lipoprotein. Data are expressed as mean ± standard deviation (n = 10). Compared with the negative group, aP < 0.05; compared with the control group, bP < 0.05; BOAA group compared with the COA group, cP < 0.05.

Figure 2 Histopathological observations of adipose tissue A: control group; B: negative group; C: met group; D: BOAA group; E: COA group; F: adipocyte size. Control group: no treatment is given; negative group: treated with high fat feed; Met group: treated with metformin; COA group: treated with COA; BOAA group: treated with BOAA. BOAA: Bo′s abdominal acupuncture; COA: conventional acupuncture. Data are expressed as mean ± standard deviation (n = 10). Compared with the control group, aP < 0.05; compared with the negative group, bP < 0.05; BOAA group compared with the COA group, cP < 0.05. Samples were stained with hematoxylin-eosin and photographed at 20 × 10 magnification.

Figure 3 Serum levels of FGF21, ADP leptin, PPAR-γ and C-P, and the mRNA expressions of FGF21, ADP, leptin, PPAR-γ, PPAR-α and AMPK in adipose tissue A: FGF21; B: ADP; C: Leptin; D: PPAR-γ; E: C-P; F-G: relative mRNA level. A-E: serum levels of FGF21, ADP leptin, PPAR-γ and C-P. F-G: q-PCR was used to analyze the mRNA expression levels of FGF21, ADP, leptin, PPAR-γ, PPAR-α and AMPK in adipose tissue. BOAA: Bo′s abdominal acupuncture; COA: conventional acupuncture; FGF21: fibroblast growth factor 21; PPAR-α: peroxisome proliferator-activated receptor α; PPAR-γ: peroxisome proliferator-activated receptor γ; AMPK: adenosine 5‘-monophosphate (AMP)-activated protein kinase; C-P: C-peptide; ADP: adiponectin. Control group: no treatment is given; negative group: treated with high fat feed; Met group: treated with metformin; COA group: treated with COA; BOAA group: treated with BOAA. Data are expressed as mean ± standard deviation (n = 10). Compared with the control group, aP < 0.05; compared with the negative group, bP < 0.05; BOAA group compared with the COA group, cP < 0.05.

Figure 4 Proteins expression of FGF21/PPAR-γ/ERK/AMPK signaling pathway in adipose tissue A-F: Western blot was used to analyze the protein levels of (B) FGF21, (C) FGFR3, (D) PPAR-α, (E) PPAR-γ, (F) ERK in adipose tissue. G-M: Western blot was used to analyze the protein levels of (H) AMPK, (I) p-AMPK, (J) LKB1, (K) p-LKB1, (L) ACC, (M) p-ACC in adipose tissue. BOAA: Bo′s abdominal acupuncture; COA: conventional acupuncture; FGF21:fibroblast growth factor 21; FGFR3:fibroblast growth factor receptors3; PPAR-α:peroxisome proliferator-activated receptor α; PPAR-γ: peroxisome proliferator-activated receptor γ; ERK: extracellular signal-regulated kinase; AMPK: adenosine 5‘-monophosphate-activated protein kinase; p-AMPK: phosphorylated adenosine 5‘-monophosphate-activated protein kinase; LKB1: liver kinase B1; p-LKB1: phosphorylated liver kinase B1; ACC: acetyl-CoA carboxylase; p-ACC: phosphorylated acetyl-CoA carboxylase. Control group: no treatment is given; negative group: treated with high fat feed; Met group: treated with metformin; COA group: treated with COA; BOAA group: treated with BOAA. Data are expressed as mean ± standard deviation (n = 10). Compared with the control group, aP < 0.05; compared with the negative group, bP < 0.05; BOAA group compared with the COA group, cP < 0.05.

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