Exploring the therapeutic potential of Morus alba Linne extract in targeting localized adiposity

doi:10.19852/j.cnki.jtcm.2025.05.004

Abstract

Abstract:

OBJECTIVE: To determine direct targeting of localized adiposity through Morus alba Linne bark injection based on pharmacology network analysis.

METHODS: Male C57BL/6J mice were fed a high-fat diet (HFD) to induce obesity. After 6 weeks on HFD, the water extract of Morus alba L.bark (MAB, 2 mg/mL) was locally injected into one inguinal fat pad, while saline was injected into the other side, 3 times/week for 6 weeks (n = 6/group). The water extract of MAB was freeze-dried and then diluted in saline before use.

RESULTS: HFD-fed mice treated with local MAB topical injection showed reduced adipocyte weight and size in inguinal fat pads by dual-energy X-ray absorptiometry. No toxicity changes seen in liver, spleen, kidney tissue, or alanine aminotransferase / aspartate aminotransferase levels in serum by MAB injection. Protein levels of phosphorylated insulin receptor substrate-1 and glucose transporter type 4, and mRNA expression of adiponectin, were increased in inguinal adipose tissue injected with MAB locally. Locally MAB injection led to a decrease in glucose-6-phosphatase and phosphoenolpyruvate carboxykinase, linked to gluconeogenesis, while forkhead box protein O1, which regulates these factors, was increased. Moreover, there was an increase in adenosine 5‘-monophosphate-activated protein kinase, related to lipogenesis, as well as elevated levels of hormone-sensitive lipase and fatty acid synthase, both associated with lipolysis. These results support the 'insulin signaling pathway' and 'regulation of lipolysis in adipocytes' identified in the Kyoto Encyclopedia of Genes and Genomes pathway through network analysis.

CONCLUSION: This study suggests that MAB topical injection exhibits localized fat reduction by inhibiting insulin resistance, gluconeogenesis and lipogenesis mediator, while activating lipolysis enzymes within targeted adipose site.

Key words: Morus Alba Linne Bark, localized fat, adiposity, obesity, network pharmacology

CHOI You Yeon, JIN Seong chul, KIM Mi Hye, BAEK Hee Kyung, KIM Dong Hyun, OH Sung Hyuk, YANG Woong Mo. Exploring the therapeutic potential of Morus alba Linne extract in targeting localized adiposity[J]. Journal of Traditional Chinese Medicine, 2025, 45(5): 970-978.

Figures/Tables 4

Table 1 MAB target pathway based on GO process and KEGG 2021 human pathway

Category	Description	P value (< 0.05)	FDR value	Background genes	Common genes
GO biological process	Cellular response to insulin stimulus	3.8E-18	1.3E-16	169	31
	Regulation of cellular response to insulin stimulus	4.0E-10	6.6E-09	72	15
	Regulation of insulin secretion	2.5E-09	3.6E-08	182	21
	Regulation of insulin receptor signalling pathway	7.9E-09	1.1E-07	64	13
	Insulin receptor signalling pathway	2.9E-08	3.7E-07	87	14
KEGG pathways	Insulin signalling pathway	9.3E-22	7.8E-21	133	32
	Regulation of lipolysis in adipocytes	1.3E-09	3.7E-09	54	13
	Insulin secretion	1.3E-04	2.4E-04	82	9

Figure 1 Effect of MAB on weight of inguinal fat tissues and the size of adipocytes in high-fat diet-induced obese mice A: weekly body weight changes in HFD-fed mice treated with MAB or saline. HFD: high-fat diet group, HFD + MAB injection: high-fat diet with MAB injection; B: gross morphology of inguinal fat pads after 18 injections of saline or MAB (100 μL per injection). B1: representative image of mouse dissection. White dotted line indicates the area of inguinal fat pads in the saline- and MAB-treated groups. scale bar = 0.5 cm, B2: quantification of inguinal fat weight relative to body weight; C: dual-energy X-ray absorptiometry analysis of body after 18 injections of saline or MAB (100 μL per injection). saline: treated with saline; MAB: treated with MAB. C1: representative dual-energy X-ray absorptiometry images showing fat (red), lean tissue (blue), and bone (white), Scale bar is 0.5 cm, C2: relative fat mass in the inguinal region after 18 injections of saline or MAB; D: histological assessment via hematoxylin and eosin staining visualized the reduction in adipocyte size after 18 injections of saline or MAB. D1: treated with saline, D2: treated with MAB; D3: quantification of average adipocyte diameter in saline- and MAB-treated groups. D4: fat diameter ratio in saline- and MAB-treated groups. Saline: treated with saline; MAB: treated with MAB. MAB: the water extract of Morus alba L. bark; HFD: high-fat diet; ANOVA: analysis of variance. Statistical significance was assessed using one-way ANOVA. Results are presented as mean ± standard deviation (n = 6). aP < 0.001, bP < 0.01, vs saline group.

Figure 2 Effect of MAB on the expression of insulin signaling-related mediators in inguinal fat tissues of obese mice A: immunofluorescence detection of GLUT4 (upper panels, red) and phospho-IRS-1 (lower panels, green) in inguinal fat pads after 18 injections of saline or MAB (100 μL per injection). Fluorescence intensity was quantified in arbitrary units. A1: GLUT4 expression in the saline-treated group; A2: GLUT4 expression in the MAB-treated group; A3: phospho-IRS-1 expression in the saline-treated group; A4: phospho-IRS-1 expression in the MAB-treated group. Scale bar = 50 μm. A5: relative fluorescence intensity of GLUT4; A6: relative fluorescence intensity of phospho-IRS-1; B: protein expression of p-IRS-1 levels in inguinal fat pad after 18 injections of saline or MAB (100 μL per injection); B1: protein expression of phospho-IRS-1 detected by Western blot; B2: quantified relative intensity of phospho-IRS-1 normalized to the saline group, which was assigned a value of 1; C: mRNA expression of adiponectin in inguinal fat pad after 18 injections of saline or MAB (100 μL per injection); C1: mRNA expression of adiponectin detected by RT-PCR; C2: quantified relative intensity of adiponectin normalized to the saline group, which was assigned a value of 1. Saline: treated with saline; MAB: treated with MAB. MAB: the water extract of Morus alba L. bark; GLUT4: glucose transporter type 4; p-IRS-1: phosphorylated insulin receptor substrate-1; ANOVA: analysis of variance. Statistical significance was assessed using one-way ANOVA. Results are presented as mean ± standard deviation (n = 6). aP < 0.001, vs saline group.

Figure 3 Effect of MAB on the expression of gluconeogenesis-mediators and AMPK-mediated FAS/HSL pathway in inguinal fat tissues of obese mice A: protein expression of gluconeogenesis-related markers in the inguinal fat pad following 18 injections of saline or MAB. A1: Western blot images showing the expression levels of FoxO1, G6Pase, and PEPCK in the inguinal fat pad of saline- and MAB-treated groups. GAPDH was used as a loading control; A2: relative intensity of FoxO1; A3: relative intensity of G6Pase; A4: relative intensity of PEPCK; B: protein expression of lipid metabolism-related markers in the inguinal fat pad following 18 injections of saline or MAB; B1: western blot images showing the expression of p-AMPKα, total AMPKα, FAS, and HSL in the inguinal fat pad; B2: relative intensity of p-AMPKα. AMPKα was used as a loading control; B3: relative intensity of FAS; B4: relative intensity of HSL. β-actin was used as a loading control. Protein expression was quantified as a relative value by normalizing it to the saline group, which was assigned a value of 1. Saline: treated with saline; MAB: treated with MAB. MAB: the water extract of Morus alba L. bark; FoxO1: forkhead box protein O1; G6Pase: glucose-6-phosphatase; PEPCK: phosphoenolpyruvate carboxykinase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; AMPK: adenosine 5‘-monophosphate-activated protein kinase; p-AMPK: phosphorylated AMPK; FAS: fatty acid synthase; HSL: hormone-sensitive lipase; ANOVA: analysis of variance. Results are presented as mean ± standard deviation (n = 6). Statistical significance was assessed using one-way ANOVA. aP < 0.001, b P < 0.01, vs saline group.

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