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. 2021 Oct 11;6(41):26959–26968. doi: 10.1021/acsomega.1c03372

Effects of Root Extract of Morinda officinalis in Mice with High-Fat-Diet/Streptozotocin-Induced Diabetes and C2C12 Myoblast Differentiation

Piao Wang †, Yi Liu †, Tong Zhang †, Cheng Yin †, Seok Yong Kang ‡, Su Jin Kim §, Yong-Ki Park †,‡,*, Hyo Won Jung †,‡,*
PMCID: PMC8529596  PMID: 34693116

Abstract

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Type 2 diabetes is the most common type of diabetes and causes a decline in muscle quality. In this study, we investigated the effects of the root extract of Morinda officinalis (MORE) on skeletal muscle damage in mice with high-fat-diet (HFD)/streptozotocin (STZ)-induced diabetes and the expression of myogenic and biogenesis regulatory proteins in C2C12 myoblast differentiation. An in vivo model comprised C57BL/6N mice fed HFD for 8 weeks, followed by a single injection of STZ at 120 mg/kg. MORE was administered at 100 and 200 mg/kg once daily (p.o.) for 4 weeks. The changes in body weight, calorie intake, and serum levels of glucose, insulin, total cholesterol (TCHO), HDL-cholesterol (HDL-C), LDL-cholesterol (LDL-C), aspartate transaminase (AST), and alanine aminotransferase (ALT) were investigated in diabetic mice. The histological changes in the gastrocnemius muscle were observed by H&E staining, and then the myofiber size was measured. The expression of the myogenic (MHC, myogenin, and MyoD) and biogenesis (PGC-1α, SIRT1, NRF1, and TFAM) regulatory proteins was examined in the muscle tissues and differentiated C2C12 myoblasts by Western blot, respectively. The administration of MORE at 200 mg/kg in mice with HFD/STZ-induced diabetes significantly reduced weight gains, calorie intake, insulin resistance, and serum levels of glucose, TCHO, LDL-C, AST, and ALT. MORE administration at 100 and 200 mg/kg significantly increased serum insulin and HDL-C levels in diabetic mice. In addition, MORE significantly increased the expression of MHC, myogenin, MyoD, PGC-1α, SIRT1, NRF1, and TFAM in muscle tissues as well as increased the myofiber size in diabetic mice. In C2C12 myoblast differentiation, MORE treatment at 0.5, 1, and 2 mg/mL significantly increased the expression of myogenic and biogenesis regulatory proteins in a dose-dependent manner. MORE improves diabetes symptoms in mice with HFD/STZ-induced diabetes by improving muscle function. This suggests that MORE could be used to prevent or treat diabetes along with muscle disorders.

Introduction

Metabolic syndromes with abdominal obesity, high blood pressure, high blood glucose,1 and high serum triglyceride levels seriously affect the regulation of insulin in glucose, fat, and protein metabolism and ultimately promote the occurrence and development of type 2 diabetes mellitus (T2DM).2,3 The incidence of T2DM with insulin resistance and deficits in the islet cell function and quantity increases with age, leading to a progressive decline in skeletal muscle mass and function4 from increasing proteolysis and decreasing protein synthesis.5 Therefore, T2DM is an important factor in the poor quality of life of older people with many complications.

The skeletal muscle is responsible for 80% of the glucose uptake in the whole body. A decline in muscle quality leads to decreased muscle capacity, insulin sensitivity, and peripheral glucose management ability that increases the risk of diabetes.6 Therefore, it has been implicated as both a cause and consequence of diabetes; a bidirectional relationship exists between muscle atrophy and T2DM.7

T2DM is medicated mainly through diet and oral hypoglycemia medications, such as metformin and other antidiabetes drugs. Unfortunately, there is no broadly effective treatment for delaying diabetes progression.8 Over the past decade, the benefits of many herbal medicines used traditionally as diabetes therapies have been demonstrated from in vivo studies and clinical trials. Traditional Chinese medicine (TCM) and Korean medicine (TKM) are becoming increasingly popular because of their effectiveness in treating many diseases with multiple targeting abilities and low side effects.

The dry root of Morinda officinalis (Rubiaceae; Morindae Radix) is a herbal medicine with a herbological character. It is warm with a sweet, spicy taste and acts on the liver and kidneys, strengthens the muscles and bones, tones the kidney yang, and enhances the immune function.9 This herb is also used to improve the symptoms, such as impotency and spermatorrhea by a kidney deficiency, menstrual irregularities, infertility,10 rheumatism, arthralgia due to wind-damp, and osteoporosis in traditional clinics.11 Recently, the antifatigue,12 anti-inflammation,13 antidepressant,14 antioxidant,15 antiosteoporosis,16 and antinociceptive effects17 of this herb have been reported. In addition, this herb has been reported to help strengthen the bones, increase bone mineral density,18 increase adipocyte differentiation,19 and have antidiabetic activity in streptozotocin (STZ)-induced diabetic rats.20

In this study, the improved effects of MORE on T2DM and its mechanism on muscle damage were investigated in HFD/STZ-induced diabetes mice and C2C12 myoblasts.

Results and Discussion

Effects of MORE on the Physiological Changes in Diabetic Mice

The effects of MORE on diabetes symptoms were examined by measuring the changes in the physiological markers, body weight, calorie intake, serum levels of glucose, insulin, and HOMA-IR in mice with HFD/STZ-induced diabetes.

As shown in Table 1, the body weight, calorie intake, and glucose level were increased significantly (p < 0.001, respectively) in diabetic mice and the insulin level was decreased significantly (p < 0.01) compared to that in the normal (Nor) group. The administration of MORE at 100 mg/kg (p < 0.01) and 200 mg/kg (p < 0.01) and metformin (Met, p < 0.001) to the diabetic mice decreased their body weights significantly. The calorie intake was significantly lower (p < 0.001, p < 0.05, and p < 0.001) in the MORE100 (p < 0.001) and MORE200 (p < 0.05) and the Met (p < 0.001) groups compared to that in the control group.

Table 1. Effects of MORE on the Physiological Changes in Mice with HFD/STZ-Induced Diabetesa.

group body weight (g) calorie intake (kcal) serum glucose (mg/dL) serum insulin (μU/mL) HOMA-IR index
Nor 28.00 ± 1.00 74.40 ± 3.10 94.60 ± 16.89 4.78 ± 0.42 0.96 ± 0.09
control 32.33 ± 2.08### 99.56 ± 5.24### 516.20 ± 33.10### 3.19 ± 0.38## 3.31 ± 0.31###
MORE100 28.67 ± 2.08** 85.59 ± 3.02*** 461.60 ± 35.23* 4.11 ± 0.61 2.61 ± 0.47*
MORE200 28.33 ± 2.51** 90.83 ± 8.01* 403.40 ± 29.05*** 4.45 ± 1.05* 2.60 ± 0.73*
Met 26.33 ± 1.52*** 76.85 ± 8.00*** 286.00 ± 37.58*** 4.68 ± 1.08** 2.43 ± 0.67**
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a

The data are expressed as the mean ± S.D. (n = 5). Nor, normal group; Control, group with HFD/STZ-induced diabetes; MORE100, mice administered 100 mg/kg MORE; mice administered 200 mg/kg MORE; and Met, group administered 250 mg/kg metformin. ##p < 0.01, ###p < 0.001 vs Nor; *p < 0.05, **p < 0.01, ***p < 0.001 vs control.

The glucose level in the serum was decreased significantly in the MORE100 (p < 0.05), MORE200 (p < 0.001), and Met (p < 0.001) groups. The insulin level was significantly higher in the MORE200 (p < 0.05) and Met (p < 0.01) groups than that in the control. On the other hand, there was no significant difference between the control group and the MORE100 group.

The HOMA-IR index showed that the insulin resistance score was significantly (p < 0.001) increased in the control group compared with that in the normal group. The insulin resistance was significantly decreased in the MORE100 and MORE200 (p < 0.05, respectively) and met (p < 0.01) groups compared with that in the control group.

Effects of MORE on the Changes in Lipid Metabolites in Diabetic Mice

The effects of MORE on the change in the lipid profile were examined by measuring the serum levels of lipid metabolites in mice with HFD/STZ-induced diabetes. As shown in Figure 1, the total cholesterol (TCHO; Figure 1A) and LDL-cholesterol (LDL-C; Figure 1B) levels were higher in the control group (p < 0.001, respectively). The MORE100 (p < 0.01) and MORE200 (p < 0.05) or Met (p < 0.01) groups had significantly decreased levels of total cholesterol (TCHO) and LDL-cholesterol (LDL-C). The HDL-cholesterol (HDL-C; Figure 1C) level in the control group was lower than that in the normal group (p < 0.05), and the levels increased after treatment with MORE or metformin (p < 0.001, respectively).

Figure 1.

Figure 1

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Effects of MORE on the serological changes in mice with HFD/STZ-induced diabetes. The levels of (A) total cholesterol (TCHO), (B) LDL-cholesterol (LDL-C), (C) HDL-cholesterol (HDL-C), (D) AST, and (E) ALT were measured in the sera of mice. The data are expressed as the mean ± S.D. (n = 5). Nor, normal group; Control, HFD/STZ-induced diabetes group; MORE100, group administered 100 mg/kg MORE; MORE200, group administered 200 mg/kg MORE; and Met, group administered 250 mg/kg metformin. #p < 0.05 and ###p < 0.001 vs Nor; *p < 0.05, **p < 0.01, and ***p < 0.001 vs Control.

The levels of AST (Figure 1D) and ALT (Figure 1E) were measured as markers of liver damage. As a result, they were increased significantly in the control group (p < 0.001, respectively) and reduced significantly in MORE100 and MORE200 or Met groups (p < 0.001, respectively).

Effects of MORE on the Histological Change in the Muscle Tissues in Diabetic Mice

The morphological changes in the muscle tissues in mice with HFD/STZ-induced diabetes were observed to investigate the effects of MORE on muscle weakness.

The main histological feature of skeletal muscle atrophy is a reduction in muscle fiber diameter and areas with a loose arrangement of muscle fibers (MFs).21 The muscle fibers of the gastrocnemius tissues in the normal group had no degradation or necrosis according to H&E staining (Figure 2A). In the diabetic mice, each bundle of muscle fiber was separated farther from others, and there was considerable space between the muscle bundles.22 The myofibers were often rounded to angular.23 On the other hand, the MORE100 and MORE200 or Met mice showed a normal structure with a significant (p < 0.001, respectively) increase in the fibers (Figure 2B).

Figure 2.

Figure 2

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Effects of MORE on the histological changes in the skeletal muscle tissues in diabetic mice. Cross sections of gastrocnemius tissues were stained with H&E stain; the nucleus stained blue, and the cytoplasm stained red under a microscope. (A) Representative pictures of tissue staining in each group (200×). The photomicrograph of the organization of the muscle fibers (MFs) into fascicles or bundles and the space between the muscle bundles (P). (B) Cross-sectional areas of myofibers in the gastrocnemius muscle were measured and expressed as mean ± S.D. (n = 5). Nor, normal group; Control, mice with HFD/STZ-induced diabetes; MORE100, group administered 100 mg/kg MORE; MORE200, group administered 200 mg/kg MORE; and Met, group administered 250 mg/kg metformin. ###p < 0.001 vs Nor; ***p < 0.001 vs Control.

Effects of MORE on the Muscle Tissue in Mice with HFD/STZ-Induced Diabetes

The effects of MORE on the skeletal muscle were examined by measuring the expression of various regulatory proteins, such as myogenetic (MyoD, myogenin, and MHC), biogenetic (PGC-1α), and atrophy (Atrogin1) in mice with HFD/STZ-induced diabetes.

As the result (Figure 3), compared to the normal group, the diabetic mice showed a significant decrease in the expressions of MHC (p < 0.05; Figure 3B), MyoD (p < 0.05; Figure 3C), myogenin (p < 0.001; Figure 3D), and PGC-1α (p < 0.001; Figure 3E) and an increase in Atrogin1 (p < 0.001, Figure 3F) in skeletal muscle tissues. The MORE100 and MORE200 mice showed a significantly increased expressions of MHC (p < 0.01), MyoD (p < 0.001), myogenin (p < 0.05), and PGC-1α (p < 0.05) in the muscle tissues but significantly decreased Atrogin1 expression (p < 0.001).

Figure 3.

Figure 3

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Effect of MORE on the muscle tissue in the mice with HFD/STZ-induced diabetes. (A) Expressions of MHC, MyoD, myogenin, PGC-1α, and Atrogin1 were determined by Western blot. (B–F) Relative expression of each protein was normalized to β-actin. The expression of each protein was measured by Western blotting and compared with the expression of β-actin as an internal control. The data are expressed as the mean ± S.D. (n = 5). Nor, normal group; Control, groups with HFD/STZ-induced diabetes; MORE100, group administered 100 mg/kg MORE; MORE200, group administered 200 mg/kg MORE; and Met, group administered 250 mg/kg metformin. #p < 0.05, ##p < 0.01, and ###p < 0.001 vs Nor; *p < 0.05, **p < 0.01, and ***p < 0.001 vs Control.

Effects of MORE on the Expression of Myogenic Proteins in C2C12 Myoblasts

The effects of MORE on myoblast differentiation were investigated by measuring the expressions of MHC, myogenin, and MyoD.

The expressions of myogenin (Figure 4B) and MyoD (Figure 4C) were increased significantly (p < 0.001, respectively) by treatment with 2 mg/mL MORE. MORE induced the differentiation of myoblasts into myotubes with elongated and widened cylinderal shapes and multiple nuclei (Figure 4F) under microscopic observation. MORE also increased MHC (Figure 4E) expression significantly at 0.5 mg/mL (p < 0.05), 1 mg/mL (p < 0.01), and 2 mg/mL (p < 0.01) in C2C12 myoblasts. The Met-treated cells showed a significant increase in myogenin, MyoD, and MHC in C2C12 cells (p < 0.01, respectively).

Figure 4.

Figure 4

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Effects of MORE on the expression of myogenic proteins in C2C12 myoblasts. The C2C12 myoblasts were differentiated into myotubes in DMEM with 2% horse serum and then treated with 0.5, 1, and 2 mg/mL MORE or 2.5 mM/mL Met for 24 h. The expressions of (A) myogenin and MyoD and (D) MHC was determined by Western blot. (B, C, E) Relative expression of each protein was normalized to β-actin. All data were presented as the mean ± SD of three independent experiments. (F) Cells were stained with anti-MHC antibody and DAPI and observed by fluorescence microscopy (200×). Green, MHC-positive cells; blue, DAPI-positive nuclei. Nor, nontreated normal cells; MORE0.5, cells treated with 0.5 mg/mL MORE; MORE1, cells treated with 1 mg/mL MORE; MORE2, cells treated with 2 mg/mL MORE; and Met, cells treated with 2.5 mM/mL metformin. #p < 0.05, ##p < 0.01, and ###p < 0.001 vs Nor.

Effects of MORE on the Expression of Biogenetic Factors in C2C12 Myoblasts

The effects of MORE on biogenesis in muscle cells were examined by measuring the expression of biogenetic factors PGC-1α, NRF1, SIRT1, and TFAM in C2C12 myotubes by Western blotting.

As shown in Figure 5, the treatment with 0.5 mg/mL MORE (p < 0.05), 1 mg/mL MORE (p < 0.01), and 2 mg/mL MORE (p < 0.001) significantly increased the expressions of PGC-1α (Figure 5B), NRF1 (p < 0.05 for 1 and 2 mg/mL; Figure 5C), SIRT1 (p < 0.05 for 0.5 and 2 mg/mL, p < 0.01 for 1 mg/mL; Figure 5D), and TFAM (p < 0.05 for 0.5 mg/mL and p < 0.01 for 1 and 2 mg/mL, Figure 5E). Met also increased the expressions of PGC-1α (p < 0.05), NRF1 (p < 0.01), SIRT1 (p < 0.05), and TFAM (p < 0.01) in C2C12 myotubes.

Figure 5.

Figure 5

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Effects of MORE on the expressions of biogenesis-regulating factors in C2C12 myoblasts. The C2C12 myoblasts were differentiated into myotubes in DMEM with 2% horse serum medium and then treated with 0.5, 1, and 2 mg/mL MORE or 2.5 mM/mL Met for 24 h. (A) Expression of TFAM, NRF1, SIRT1, and TFAM was determined by Western blot. (B–E) Relative expression of each protein was normalized to β-actin. All data are presented as the mean ± SD of three independent experiments (n = 3). Nor, nontreated normal cells; MORE0.5, cells treated with 0.5 mg/mL MORE; MORE1, cells treated with 1 mg/mL MORE; MORE2, cells treated with 2 mg/mL MORE; and Met, cells treated with 2.5 mM/mL metformin. #p < 0.05, ##p < 0.01, and ###p < 0.001 vs Nor.

Type 2 diabetes (T2DM) is the most common type of diabetes characterized by hyperglycemia and urine sugar resulting from resistance to insulin action and an inadequate compensatory insulin secretory response.24 Approximately 90% of individuals with obesity and the elderly have a high incidence of diabetes,25 and the disease is growing rapidly in developing countries.26 Medications for diabetes patients include various drugs, such as metformin, insulin-releasing pills, starch blockers, amylin analogues, and incretin-based therapeutics for lower blood sugar.27 In the concepts of traditional medicines, the human body includes two aspects, blood (Yin) and Qi (Yang). The Yin-blood is the material basis for motivation and Yang-qi is the power of the human body.28 Type 2 diabetes is thought to be caused by deficiency syndromes, which mainly include Qi deficiency, Yin deficiency, and Yang deficiency,29 and dietary irregularities, emotional disharmony, and phlegm wet constitution (obesity), which lead to three main diabetes symptoms of polydipsia, polyuria, and polyphagia.30,31

Type 2 diabetes involves insulin resistance, which could stimulate glucose uptake, severely change energy metabolism, and negatively affect skeletal muscle quality through protein degradation and mitochondrial dysfunction.32,33 Thus, insulin resistance is regarded as an important contributor to decreased muscle growth and development and induced fiber atrophy in diabetes myopathy. Skeletal muscle also is a key factor in the development of insulin resistance because quantitatively it is the principal tissue that contributes to whole-body insulin-mediated glucose disposal. Therefore, it is believed that diabetes is associated with excessive muscle loss (atrophy) that contributes to functional abnormalities of muscle.34 Recently, it has been reported that sarcopenia, a type of muscle atrophy occurring in aging, is induced by poor glucose disposal and a decreased metabolic rate and physical activity that is considered to increase the risk of developing T2DM in the elderly.35,36 At this point, it is being studied that antidiabetes drugs such as metformin may alter the balance between protein synthesis and degradation through various regulation mechanisms of muscle mass and improve muscle function by controlling various regulatory factors on myogenesis and mitochondrial biogenesis.37,38 However, the prevention of muscle loss and dysfunction is viewed as being crucial in terms of treating T2DM. In our study, we investigated the effects of MO, a herbal medicine used for strengthening the muscles and increasing energy in the elderly, on muscle function in mice with HFD/STZ-induced diabetes.

A high-calorie diet and obesity are the main risk factors for T2DM.39,40 In the present study, HFD-fed mice showed an increase in body weight, induction of both insulin resistance, and glucose tolerance that may mimic the human lifestyle.41−43 Streptozotocin is selectively toxic to the insulin-producing β cells of the pancreas in mammals and has been long injected in experimental animals for the reproduction of diabetes pathology in humans.44 Multiple methods of STZ dosing depend on the type and the severity of diabetes in mice, and the dose range also varies largely, so they are applied in various ways from 100 to 200 mg/kg in a single high dose. The mice with HFD/STZ-induced diabetes are a useful experimental animal model for screening hypoglycemic drugs and examining the corresponding mechanisms.45 In this study, we produced a diabetes mouse model with an HFD for 12 weeks, and a single injection of STZ at 120 mg/kg, a single moderately sized dose could then induced diabetes symptoms. Our diabetes model could not observe dead mice during drug administration. We also used two different dosages of MORE at 100 and 200 mg/kg according to the previous study.18,20

The main characteristics of human T2DM are high blood glucose, increase in body weight, more food/calorie intake, and lipid metabolism disorders in a clinical setting.46,47 Our in vivo model exhibited these symptoms, which were improved by MORE administration for 4 weeks. This suggests that MORE may improve the symptoms of diabetes such as obesity, insulin resistance, and T2DM, which have been associated with muscle atrophy. Although there are few studies of MO efficacy in T2DM or muscle function, it is reported that the ethanol extract of MO at 150 mg/kg significantly reduced fasting serum glucose levels in rats with STZ-induced diabetes;20 interestingly, the stem bark extract of M. lucida, the sister species of MO, has reducing effects at 500 mg/kg on increases in weight loss, water and food consumption, lipid peroxidation, and cholesterol, triglycerides, and glucose in rats with STZ-induced diabetes.48 In this study, we compared the efficacies of MORE and metformin as the first medicine to treat T2DM, which decreases hepatic glucose production and increases the insulin sensitivity of target cells. Administration of metformin at 250 mg/kg in mice with HFD/STZ-induced diabetes was shown to have similar effects on physiological and serological changes as MORE.

Dyslipidemia is believed to be a major risk factor for the development of various diabetes complications and is related mainly to changes in serum lipoproteins with high levels of TCHO, LDL-C, AST, and ALT, and a low level of HDL-C.49 In our study, an increase in serum levels of TCHO, LDL-C, AST, and ALT and a low level of HDL-C were observed in the mice with HFD/STZ-induced diabetes compared to the normal group. These levels were reduced in the MORE100 and MORE200 groups, suggesting that MORE can improve dyslipidemia in diabetes. Our finding that the TCHO level is reduced is consistent with previous reports.20

Previous studies explained that dyslipidemia leads to the apoptosis and atrophy of skeletal muscle.50 The present study observed pathological changes to the gastrocnemius muscle tissues of mice. As a result, the diabetic mice showed histological changes in the muscle tissues, such as a reduction in myofiber size with a loose arrangement and decreasing numbers. On the other hand, the diabetic mice administered MORE at low and high doses showed similar structures to the normal group. This result indicates that MORE can decrease the glucose level, reduce abnormal lipid metabolism, and decrease muscle atrophy finally in the mice with HFD/STZ-induced diabetes.

Differentiation of the skeletal muscle is a dynamic multistep process that involves multiple myogenic regulatory factors, such as MyoD, myogenin, and MHC.51 MyoD plays a crucial role in the differentiation of satellite cells into myoblasts.52 Myogenin controls the differentiation of myoblasts to myotubes53 and MHC is a differentiation marker in neo-formed myotubes.54,55 In our study, it was observed that the decreased expression of myogenic proteins in muscle tissues of diabetic mice which were increased by MORE administration. One of the features of diabetes is an altered mitochondrial function in skeletal muscle. Thus, impaired mitochondrial glycolytic capacity and altered β-oxidation in the muscle of T2DM patients have been reported.56 The AMPK/SIRT1/PGC-1α signaling pathways on energy homeostasis in muscle under diabetes conditions have been studied.57 The mitochondria are essential organelles for ATP production and regulate the lipid and glucose metabolism in the skeletal muscle.58 PGC-1α, a metabolic mediator has a crucial role in the regulation of mitochondrial biogenesis by co-activating NRF1,59 which subsequently increases the expression of TFAM.60 SIRT1 is another crucial factor of glucose and lipid metabolism and stimulates mitochondrial biogenesis via the PGC-1α/AMPK pathway.61,62 Therefore, muscle differentiation and mitochondrial biogenesis are associated with an increase in muscle mass and strength. In our study, MORE significantly increased the expression of myogenic proteins and biogenesis regulatory proteins in muscle tissues of not only diabetic mice but also in C2C12 myotubes. Moreover, a comparison of the effects of MORE at 2 mg/mL and metformin at 2.5 mM51 showed that MORE was more effective in the expression of MHC, MyoD, myogenin, and PGC-1α than metformin. This suggests that MORE can be more effective in enhancing muscle function than metformin.

MORE is contains various ingredients such as oligosaccharides, polysaccharides, iridoid glucosides, plant sterols, anthraquinones, flavonoids, vitamin C, and other ingredients,63,64 but their effects on obesity, diabetes, and muscle wasting. However, the components of MORE need to be considered in a future study.

Conclusions

The administration of MORE at 100 and 200 mg/kg for 4 weeks in mice with HFD/STZ-induced diabetes resulted in decreased body weight, calorie intake, high glucose and lipid levels, and increased insulin levels. MORE improved the mass loss of the skeletal muscle by enhancing the expression of myogenetic proteins, MyoD, myogenin, MHC and biogenetic proteins, PGC-1α, NRF1, SIRT1, and TFAM in the skeletal muscle tissues of diabetic mice and C2C12 myoblasts differentiation. These results suggest that MORE can be a preventative and therapeutic natural drug for T2DM patients showing muscle loss.

Experimental Section

Preparation of MORE

The dried root of M. officinalis (Morindae Radix) was purchased from a medicinal materials company (Kwangmyungdang Herbal Company, Ulsan, Korea) and identified by a botanical expert, Dr. Y.-K. Park, College of Korean Medicine, Dongguk University, Korea. Approximately 200 g of small pieces of M. officinalis was boiled with 2000 mL of distilled water in a round-bottom flask under reflux and extracted twice for 3 h. The crude extract was filtrated through Whatman Filter Paper (GE Healthcare UK Limited, Buckinghamshire, U.K.), concentrated under a vacuum rotary evaporator (Eyela Co. Ltd., Tokyo, Japan) at 60 °C, and then lyophilized in a freeze-dryer (IlShin Lab Co., Yangju, Korea) at −80 °C under 5 mTorr. The powder of the MORE (yield = 58%) was stored at −20 °C until needed in in vivo and in vitro study.

Preparation of In Vivo Model

Male C57BL/6N mice (Koatech, Pyeongtaek, Korea) were used in this study. The Institutional Animal Care and Use Committee of the Dongguk University (IACUC-2018-11) approved the animal care protocol and experimental procedures. The mice were acclimated to the feeding protocol 1 week before the experiment. The mice were housed in cages at 22 ± 3 °C and relative humidity of 60 ± 5% and a 12/12 h light/dark cycle. They were given access to food and water ad libitum. The mice were divided randomly into two groups: a normal group (Nor) with standard chow [3.10 kcal/g, 18% kcal from fat, 58% kcal from carbohydrates, and 24% kcal from protein (Cat. 2018S, Envigo, IN)] and a group with a high-fat diet (HFD, 5.24 kcal/g, 60% kcal from fat, and 20% kcal from carbohydrates, 20% kcal from protein) (Cat. D12492, Research diets, NJ) for 8 weeks. After feeding, the HFD mice were single injected intraperitoneally (i.p.) with STZ at 120 mg/kg body weight (b.w.), which was dissolved in citrate buffer (pH 4.5). To confirm the induction of diabetes, the fasting blood glucose (FBG) levels were measured in the whole blood of the tails using an Accu-Chek Inform II Glucose Meter (Roche, Basel, Swiss). Mice with a blood glucose concentration of more than 300 mg/dL were used.

The mice with HFD/STZ-induced were divided randomly into four groups (n = 5 per each group): the control group with HFD/STZ-induced diabetes (control), group administered MORE at 100 mg/kg b.w. (MORE100, p.o.) and 200 mg/kg b.w. (MORE200, p.o.) and that adminstered metformin at 250 mg/kg b.w. (Met, p.o.) as a reference group. Mice were administered drugs once daily for 4 weeks. The normal group was fed the normal diet, and the other groups were kept on HFD during drug administration.

Measurement of Physiological Parameters

At the end of the experiment, the body weight and food intake were measured. Also, the caloric intake was calculated by food intake multiplying 3.1 kcal/g in the standard diet and multiplying 5.24 kcal/g in the HFD mice.

Measurement of Serological Parameters

The animals were sacrificed using a 5% isoflurane in O2 (75%) and N2O (25%) euthanasia station after 24 h fasting. Whole blood was harvested from the abdominal aorta using a syringe. The blood samples were isolated by centrifugation twice at 3000 rpm for 10 min at room temperature and the serum samples were isolated for serological analysis. The following were measured including levels of glucose (GLU), aspartate transaminase (AST), alanine aminotransferase (ALT), total cholesterol (TCHO), high-density lipoprotein-cholesterol (HDL-C), and low-density lipoprotein-cholesterol (LDL-C) in the serum samples using an automated clinical chemistry analyzer (FDC7000i, Fujifilm Co, Tokyo, Japan). The insulin levels were analyzed using an insulin enzyme-linked immunosorbent Assay (ELISA) kit (Cat. 90082, Crystal Chem Inc., IL) according to the manufacturer’s protocol. The concentration of insulin in the serum samples was determined using a standard curve. The equation calculated the level of homeostasis model assessment of insulin resistance (HOMA-IR) as follows: HOMA-IR = [fasting insulin (μU/mL) × fasting serum glucose (mM)]/22.5.

Histopathological Observation

The gastrocnemius tissues were obtained for histological observations and protein expression analysis. The gastrocnemius muscle tissues were immobilized with 4% paraformaldehyde for 24 h to make a specimen section after dehydration paraffin embedding and stained with hematoxylin and eosin (H&E). The change in muscle tissues was observed under a microscope (original magnification 200×). The size of fibers was measured in the image using the ImageJ software (https://imagej.nih.gov/ij/) and presented the mean cross-sectional area of the muscle fibers.65

Cell Culture and Treatment

The C2C12 cells (ATCC, VA), a mouse myoblast line, were cultured in DMEM (Corning, NY) supplemented with 10% fetal bovine serum (Merck Millipore, MA) with penicillin/streptomycin (Corning) at 37 °C in a 5% CO2 incubator. When more than 90% confluence was reached, the medium was changed to DMEM supplemented with 2% horse serum (Thermo Fisher Scientific, MA) for 4 days and then treated with MORE at different concentrations (0.5, 1, and 2 mg/mL) and metformin 2.5 mM/mL for 24 h. No toxicity was observed up to 2 mg/mL by an MTT assay (Figure S1).

Immunocytochemistry Staining

The C2C12 cells were seeded on Thermanox plastic coverslips (Nunc, Thermo Fisher Scientific) and differentiated with 2% horse serum for 4 days. After treatment with MORE or metformin for 24 h, the C2C12 myotubes were washed three times with 1× PBS, fixed in 4% paraformaldehyde, placed on ice for 20 min, and permeabilized in 0.5% Triton X-100 (Sigma-Aldrich, St. Louis, MO) for 5 min at room temperature. Subsequently, the cells were washed with 1× PBS, blocked in 0.5% bovine serum albumin (BSA) for 2 h, and incubated with the anti-MHC (Cat. Sc-376157, Santa Cruz Biotechnology Inc., TX) antibody at 4 °C overnight. The myotubes were washed three times with 1× PBS and incubated with a goat antimouse antibody conjugated with Alexa Fluor 488 (Thermo Fisher Scientific, Cat. A11001). The nuclei were counterstained with DAPI. The cells were visualized using a fluorescence microscope (Leica DM2500, Leica Microsystems, Wetzlar, Germany). The MHC-positive cells were observed by green fluorescence in blue multinucleated myotubes with DAPI.

Western Blot Analysis

To isolate the total protein, the skeletal muscle tissues and C2C12 myotubes were homogenized with a protein lysis buffer containing a protease inhibitor and phosphatase inhibitor, homogenized using a homogenizer (T10 basic, IKA, Staufen im Breis-gau, Germany), and centrifuged at 14000 rpm at 4 °C for 20 min. After measuring the total protein concentration in each sample, an equal amount of protein (30 μg) was separated into 8–10% acrylamide gels, and SDS-polyacrylamide gel electrophoresis (PAGE) was performed. The gels were transferred onto nylon membranes and blocked with 5% skim milk for 1 h at room temperature. The membranes were incubated with the specific primary antibodies against MyoD (sc-377460, Santa Cruz), MHC (sc-376157), myogenin (sc-52903), PGC-1α (Cat. NBP1-04676, Bioss Antibodies, MA), SIRT1 (Cat. bs-0921R), NRF1 (Cat. 69432s, Cell Signaling, MA), TFAM (Cat. PA5-68789, Thermo Fisher Scientific), and Atrogin1 (Cat. PA5-91959) at 4 °C overnight. The membranes were then washed three times with 1× Tris-buffered saline (pH 7.4) containing 0.1% Tween-20 (TBST) buffer for 15 min. The membranes were incubated with the secondary antibodies horseradish peroxidase (HRP)-labeled antimouse or antirabbit IgG (Bio-Rad, CA) for 2 h at room temperature and washed three times with 1× TBST buffer. The protein signals were detected using an ECL substrate Western blotting detection reagent (Bio-Rad). ChemiDoc MP Imaging System (Bio-Rad) was used to capture the image, which was quantified by densitometry using ImageJ programming software (ImageJ, NIH, MD). The expression of each target protein was normalized to β-actin (Sigma-Aldrich) as an internal control.

Statistical Analysis

All data are presented as mean ± standard deviation (mean ± SD; n = 5 per each group in vivo and n = 3 in vitro). Statistical analysis was performed using a one-way analysis of variance (ANOVA) followed by a Dunnett’s test between normal and control groups or control and drug groups. A p value <0.05 was considered significant. All statistical analyses were performed on the GraphPad Prism software Version 5.0 (GraphPad Software, CA).

Glossary

Abbreviations

MORE

Morinda officinalis root extract

T2DM

type 2 diabetes mellitus

HFD

high-fat diet

STZ

streptozotocin

TCHO

total cholesterol

HDL-C

HDL-cholesterol

LDL-C

LDL-cholesterol

AST

aspartate transaminase

ALT

alanine aminotransferase

TCM

Traditional Chinese medicine

TKM

Traditional Korean medicine

FBG

fasting blood glucose

Supporting Information Available

The Supporting Information is available free of charge at https://pubs.acs.org/doi/10.1021/acsomega.1c03372.

  • Cell viability assay: effects of MORE on the viability of C2C12 cells. The data used to support the study are available from the corresponding author upon request. (PDF)

Author Contributions

H.W.J. and Y.-K.P. designed the study. P.W., Y.L., T.Z., C.Y., and S.Y.K. performed the experiments. P.W., S.Y.K., and S.J.K. conducted the statistical analyses. P.W. and H.W.J. wrote the manuscript. S.Y.K., H.W.J., Y.-K.P., and S.J.K. revised the manuscript, and all authors approved the final version.

This research was funded by the National Research Foundation of Korea (NRF) [2016R1D1A1B04935601 and 2021R1A2C1012267].

The authors declare no competing financial interest.

Notes

All procedures were performed in accordance with the guidelines of the Dongguk University of Korea’s Medicine ethics committee.

Supplementary Material

ao1c03372_si_001.pdf (665.1KB, pdf)

References

  1. Martin B. C.; Warram J. H.; Krolewski A. S.; Soeldner J.; Kahn C.; Bergman R.; et al. Role of glucose and insulin resistance in development of type 2 diabetes mellitus: results of a 25-year follow-up study. Lancet 1992, 340, 925–929. 10.1016/0140-6736(92)92814-V. [DOI] [PubMed] [Google Scholar]
  2. Huang J.; Wang X.; Tao G.; Song Y.; Ho C.; Zheng J.; Ou S. Feruloylated oligosaccharides from maize bran alleviate the symptoms of diabetes in streptozotocin-induced type 2 diabetic rats. Food Funct. 2018, 9, 1779–1789. 10.1039/C7FO01825H. [DOI] [PubMed] [Google Scholar]
  3. Peng S.; Wei P.; Lu Q.; Liu R.; Ding Y.; Zhang J. Beneficial Effects of Poplar Buds on Hyperglycemia, Dyslipidemia, Oxidative Stress, and Inflammation in Streptozotocin-Induced Type-2 Diabetes. J. Immunol. Res. 2018, 2018, 7245956 10.1155/2018/7245956. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Akhmedov D.; Berdeaux R. The effects of obesity on skeletal muscle regeneration. Front. Physiol. 2013, 4, 371 10.3389/fphys.2013.00371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Zhang J.; Zhuang P.; Wang Y.; Song L.; Zhang M.; Lu Z.; Zhang L.; Wang J.; Alemu P. N.; Zhang Y.; et al. Reversal of muscle atrophy by Zhimu-Huangbai herb-pair via Akt/mTOR/FoxO3 signal pathway in streptozotocin-induced diabetic mice. PLoS One 2014, 9, e100918 10.1371/journal.pone.0100918. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Kim K.-S.; Park K.-S.; Kim M.-J.; Kim S.-K.; Cho Y.-W.; Park S. W. Type 2 diabetes is associated with low muscle mass in older adults. Geriatr. Gerontol. Int. 2014, 14, 115–121. 10.1111/ggi.12189. [DOI] [PubMed] [Google Scholar]
  7. Mesinovic J.; Zengin A.; De Courten B.; Ebeling P. R.; Scott D. Sarcopenia and type 2 diabetes mellitus: a bidirectional relationship. Diabetes, Metab. Syndr. Obes. 2019, 12, 1057. 10.2147/DMSO.S186600. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Chaudhury A.; Duvoor C.; Reddy Dendi V. S.; Kraleti S.; Chada A.; Ravilla R.; Marco A.; Shekhawat N. S.; Montales M. T.; Kuriakose K.; et al. Clinical review of antidiabetic drugs: implications for type 2 diabetes mellitus management. Front. Endocrinol. 2017, 8, 6 10.3389/fendo.2017.00006. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chinese Pharmacopoeia Commission . Pharmacopoeia of the Peoples Republic of China, 1st ed.; Chinese Medical Science and Technology Press: Beijing, China, 2020; pp 83–84. [Google Scholar]
  10. Chen D.-L.; Li N.; Lin L.; Long H.-m.; Lin H.; Chen J.; Zhang H.-M.; Zeng C.-c.; Liu S.-H. Confocal mirco-Raman spectroscopic analysis of the antioxidant protection mechanism of the oligosaccharides extracted from Morinda officinalis on human sperm DNA. J. Ethnopharmacol. 2014, 153, 119–124. 10.1016/j.jep.2014.01.021. [DOI] [PubMed] [Google Scholar]
  11. Jiang K.; Huang D.; Zhang D.; Wang X.; Cao H.; Zhang Q.; Yan C. Investigation of inulins from the roots of Morinda officinalis for potential therapeutic application as anti-osteoporosis agent. Int. J. Biol. Macromol. 2018, 120, 170–179. 10.1016/j.ijbiomac.2018.08.082. [DOI] [PubMed] [Google Scholar]
  12. Zhang H.-l.; Li J.; Li G.; Wang D.-m.; Zhu L.-p.; Yang D.-p. Structural characterization and anti-fatigue activity of polysaccharides from the roots of Morinda officinalis. Int. J. Biol. Macromol. 2009, 44, 257–261. 10.1016/j.ijbiomac.2008.12.010. [DOI] [PubMed] [Google Scholar]
  13. Liang J.; Liang J.; Hao H.; Lin H.; Wang P.; Wu Y.; Jiang X.; Fu C.; Li Q.; Ding P.; et al. The extracts of Morinda officinalis and its hairy roots attenuate dextran sodium sulfate-induced chronic ulcerative colitis in mice by regulating inflammation and lymphocyte apoptosis. Front. Immunol. 2017, 8, 905 10.3389/fimmu.2017.00905. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Zhang Z.-Q.; Yuan L.; Yang M.; Luo Z.-P.; Zhao Y.-M. The effect of Morinda officinalis How, a Chinese traditional medicinal plant, on the DRL 72-s schedule in rats and the forced swimming test in mice. Pharmacol., Biochem. Behav. 2002, 72, 39–43. 10.1016/S0091-3057(01)00730-4. [DOI] [PubMed] [Google Scholar]
  15. Chang M.-S.; Kim W.-N.; Yang W.-M.; Kim H.-Y.; Oh J.-H.; Park S.-K. Cytoprotective effects of Morinda officinalis against hydrogen peroxide-induced oxidative stress in Leydig TM3 cells. Asian J. Androl. 2008, 10, 667–674. 10.1111/j.1745-7262.2008.00414.x. [DOI] [PubMed] [Google Scholar]
  16. Wu Y.-B.; Zheng C.-J.; Qin L.-P.; Sun L.-N.; Han T.; Jiao L.; Zhang Q.-Y.; Wu J.-Z. Antiosteoporotic activity of anthraquinones from Morinda officinalis on osteoblasts and osteoclasts. Molecules 2009, 14, 573–583. 10.3390/molecules14010573. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kim I.-T.; Park H.-J.; Nam J.-H.; Park Y.-M.; Won J.-H.; Choi J.; Choe B.-K.; Lee K.-T. In-vitro and in-vivo anti-inflammatory and antinociceptive effects of the methanol extract of the roots of Morinda officinalis. J. Pharm. Pharmacol. 2010, 57, 607–615. 10.1211/0022357055902. [DOI] [PubMed] [Google Scholar]
  18. MengYong Z.; CaiJiao W.; HuSheng Z.; XianWu P.; JianMin F. Protective effect of polysaccharides from Morinda officinalis on bone loss in ovariectomized rats. Int. J. Biol. Macromol. 2008, 43, 276–278. 10.1016/j.ijbiomac.2008.06.008. [DOI] [PubMed] [Google Scholar]
  19. Liu Q.; Kim S. B.; Ahn J. H.; Hwang B. Y.; Kim S. Y.; Lee M. K. Anthraquinones from Morinda officinalis roots enhance adipocyte differentiation in 3T3-L1 cells. Nat. Prod. Res. 2012, 26, 1750–1754. 10.1080/14786419.2011.608676. [DOI] [PubMed] [Google Scholar]
  20. Soon Y.; Tan B. Evaluation of the hypoglycemic and anti-oxidant activities of Morinda officinalis in streptozotocin-induced diabetic rats. Singapore Med. J. 2002, 43, 077–085. [PubMed] [Google Scholar]
  21. Ma J.; Meng X.; Liu Y.; Yin C.; Zhang T.; Wang P.; Park Y.-K.; Jung H. W. Effects of a rhizome aqueous extract of Dioscorea batatas and its bioactive compound, allantoin in high fat diet and streptozotocin-induced diabetic mice and the regulation of liver, pancreas and skeletal muscle dysfunction. J. Ethnopharmacol. 2020, 259, 112926 10.1016/j.jep.2020.112926. [DOI] [PubMed] [Google Scholar]
  22. Sayed R. K.; de Leonardis E. C.; Guerrero-Martínez J. A.; Rahim I.; Mokhtar D. M.; Saleh A. M.; Abdalla K. E.; Pozo M. J.; Escames G.; López L. C.; et al. Identification of morphological markers of sarcopenia at early stage of aging in skeletal muscle of mice. Exp. Gerontol. 2016, 83, 22–30. 10.1016/j.exger.2016.07.007. [DOI] [PubMed] [Google Scholar]
  23. Berridge B. R.; Van Vleet J. F.; Herman E. In Haschek and Rousseaux’s Handbook of Toxicologic Pathology, 3rd ed.; Haschek W. M.; Rousseaux C. G.; Wallig M. A., Eds.; Academic Press: Boston, 2013; pp 1567–1665. [Google Scholar]
  24. Diagnosis and classification of diabetes mellitus. Diabetes Care 2013, 36, S67–S74. 10.2337/dc13-S067. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Salas-Salvadó J.; Martinez-Gonzalez M.; Bullo M.; Ros E. The role of diet in the prevention of type 2 diabetes. Nutr., Metab. Cardiovasc. Dis. 2011, 21, B32–B48. 10.1016/j.numecd.2011.03.009. [DOI] [PubMed] [Google Scholar]
  26. Shaw J. E.; Sicree R. A.; Zimmet P. Z. Global estimates of the prevalence of diabetes for 2010 and 2030. Diabetes Res. Clin. Pract. 2010, 87, 4–14. 10.1016/j.diabres.2009.10.007. [DOI] [PubMed] [Google Scholar]
  27. Medication for Type 2 Diabetes; Institute for Quality and Efficiency in Health Care (IQWiG): Cologne, Germany, 2006.
  28. Tang J.-L.; Liu B.-Y.; Ma K.-W. Traditional chinese medicine. Lancet 2008, 372, 1938–1940. 10.1016/S0140-6736(08)61354-9. [DOI] [PubMed] [Google Scholar]
  29. Wang J.; Ma Q.; Li Y.; Li P.; Wang M.; Wang T.; Wang C.; Wang T.; Zhao B. Research progress on traditional Chinese medicine syndromes of diabetes mellitus. Biomed. Pharmacother. 2020, 121, 109565 10.1016/j.biopha.2019.109565. [DOI] [PubMed] [Google Scholar]
  30. Yang Y.Questions and Answers in Traditional Chinese Medicine, 1st ed.; People’s Medical Publishing House: Beijing, 1985; pp 15–16. [Google Scholar]
  31. Ge J.; Xu Y.; Wang Z.. Diabetology, 2nd ed.; Shanghai Science and Technology Press: Shanghai, 2020; pp 16–18. [Google Scholar]
  32. Greene N. P.; Brown J. L.; Rosa-Caldwell M. E.; Lee D. E.; Blackwell T. A.; Washington T. A. Skeletal muscle insulin resistance as a precursor to Diabetes: Beyond glucoregulation. Curr. Diabetes Rev. 2018, 14, 113–128. 10.2174/1573399813666161122123636. [DOI] [PubMed] [Google Scholar]
  33. Sala D.; Zorzano A. In Molecular Nutrition and Diabetes, 1st ed; Mauricio D., Ed.; Academic Press: San Diego, 2016; pp 73–85. [Google Scholar]
  34. Kautzky-Willer A.; Harreiter J.; Pacini G. Sex and gender differences in risk, pathophysiology and complications of type 2 diabetes mellitus. Endocr. Rev. 2016, 37, 278–316. 10.1210/er.2015-1137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Park S. W.; Goodpaster B. H.; Strotmeyer E. S.; de Rekeneire N.; Harris T. B.; Schwartz A. V.; Tylavsky F. A.; Newman A. B. Decreased muscle strength and quality in older adults with type 2 diabetes: the health, aging, and body composition study. Diabetes 2006, 55, 1813–1818. 10.2337/db05-1183. [DOI] [PubMed] [Google Scholar]
  36. Cetrone M.; Mele A.; Tricarico D. Effects of the antidiabetic drugs on the age-related atrophy and sarcopenia associated with diabetes type II. Curr. Diabetes Rev. 2014, 10, 231–237. 10.2174/1573399810666140918121022. [DOI] [PubMed] [Google Scholar]
  37. Wu C.-N.; Tien K.-J. The Impact of Antidiabetic Agents on Sarcopenia in Type 2 Diabetes: A Literature Review. J. Diabetes Res. 2020, 2020, 9368583 10.1155/2020/9368583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Han S. E.; Kim S. J.; Kim Y. I.; Nam-Goong I. S.; Jung H. W.; Kim E. S. Enhancing effects of anagliptin on myoblast differentiation and the expression of mitochondrial biogenetic factors in C2C12 mouse skeletal muscle cells. Clin. Exp. Pharmacol. Physiol. 2020, 47, 903–906. 10.1111/1440-1681.13255. [DOI] [PubMed] [Google Scholar]
  39. Yajnik C. S. The insulin resistance epidemic in India: fetal origins, later lifestyle, or both. Nutr. Rev. 2001, 59, 1–9. 10.1111/j.1753-4887.2001.tb01898.x. [DOI] [PubMed] [Google Scholar]
  40. Yoon K.-H.; Lee J.-H.; Kim J.-W.; Cho J. H.; Choi Y.-H.; Ko S.-H.; Zimmet P.; Son H.-Y. Epidemic obesity and type 2 diabetes in Asia. Lancet 2006, 368, 1681–1688. 10.1016/S0140-6736(06)69703-1. [DOI] [PubMed] [Google Scholar]
  41. Lv Y.; Ren W.; Zhang Y.; Huang Y.; Hao J.; Ma K.; Ma Y.; Yang X. Antidiabetic effects of a lipophilic extract obtained from flowers of Wisteria sinensis by activating Akt/GLUT4 and Akt/GSK3β. Food Nutr. Res. 2020, 64, 3589 10.29219/fnr.v64.3589. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Li W.; Zhu C.; Liu T.; Zhang W.; Liu X.; Li P.; Zhu T.. Epigallocatechin-3-gallate ameliorates glucolipid metabolism and oxidative stress in type 2 diabetic rats Diabetes Vasc. Dis. Res. 202017, 1−10 10.1177/1479164120966998. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Al-Trad B.; Alkhateeb H.; Alsmadi W.; Al-Zoubi M. Eugenol ameliorates insulin resistance, oxidative stress and inflammation in high fat-diet/streptozotocin-induced diabetic rat. Life Sci. 2019, 216, 183–188. 10.1016/j.lfs.2018.11.034. [DOI] [PubMed] [Google Scholar]
  44. Deeds M.; Anderson J.; Armstrong A.; Gastineau D.; Hiddinga H.; Jahangir A.; Eberhardt N.; Kudva Y. Single dose streptozotocin-induced diabetes: considerations for study design in islet transplantation models. Lab. Anim. 2011, 45, 131–140. 10.1258/la.2010.010090. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Asrafuzzaman M.; Cao Y.; Afroz R.; Kamato D.; Gray S.; Little P. J. Animal models for assessing the impact of natural products on the aetiology and metabolic pathophysiology of Type 2 diabetes. Biomed. Pharmacother. 2017, 89, 1242–1251. 10.1016/j.biopha.2017.03.010. [DOI] [PubMed] [Google Scholar]
  46. Han H. S.; Jeon H.; Kang S. C. Phellopterin isolated from Angelica dahurica reduces blood glucose level in diabetic mice. Heliyon 2018, 4, e00577 10.1016/j.heliyon.2018.e00577. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Sajak A. A. B.; Mediani A.; Ismail A.; Abas F.; et al. Metabolite Variation in Lean and Obese Streptozotocin (STZ)-Induced Diabetic Rats via 1H NMR-Based Metabolomics Approach. Appl. Biochem. Biotechnol. 2017, 182, 653–668. 10.1007/s12010-016-2352-9. [DOI] [PubMed] [Google Scholar]
  48. Domekouo U. L.; Longo F.; Tarkang P. A.; Tchinda A. T.; Tsabang N.; Donfagsiteli N. T.; Tamze V.; Kamtchouing P.; Agbor G. A. Evaluation of the antidiabetic and antioxidant properties of Morinda lucida stem bark extract in streptozotocin intoxicated rats. Pak. J. Pharm. Sci. 2016, 29, 903–911. [PubMed] [Google Scholar]
  49. Mooradian A. D. Dyslipidemia in type 2 diabetes mellitus. Nat. Rev. Endocrinol. 2009, 5, 150–159. 10.1038/ncpendmet1066. [DOI] [PubMed] [Google Scholar]
  50. Sishi B.; Loos B.; Ellis B.; Smith W.; du Toit E. F.; Engelbrecht A.-M. Diet-induced obesity alters signalling pathways and induces atrophy and apoptosis in skeletal muscle in a prediabetic rat model. Exp. Physiol. 2011, 96, 179–193. 10.1113/expphysiol.2010.054189. [DOI] [PubMed] [Google Scholar]
  51. Ma J.; Kang S. Y.; Meng X.; Kang A. N.; Park J. H.; Park Y.-K.; Jung H. W. Effects of rhizome extract of dioscorea batatas and its active compound, allantoin, on the regulation of myoblast differentiation and mitochondrial biogenesis in c2c12 myotubes. Molecules 2018, 23, 2023 10.3390/molecules23082023. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Smith C.; Janney M.; Allen R. Temporal expression of myogenic regulatory genes during activation, proliferation, and differentiation of rat skeletal muscle satellite cells. J. Cell. Physiol. 1994, 159, 379–385. 10.1002/jcp.1041590222. [DOI] [PubMed] [Google Scholar]
  53. Nabeshima Y.; Hanaoka K.; Hayasaka M.; Esuml E.; Li S.; Nonaka I.; Nabeshima Y.-i. Myogenin gene disruption results in perinatal lethality because of severe muscle defect. Nature 1993, 364, 532–535. 10.1038/364532a0. [DOI] [PubMed] [Google Scholar]
  54. Dedieu S.; Mazeres G.; Cottin P.; Brustis J.-J. Involvement of myogenic regulator factors during fusion in the cell line C2C12. Int. J. Dev. Biol. 2002, 46, 235–241. [PubMed] [Google Scholar]
  55. Sartorelli V.; Caretti G. Mechanisms underlying the transcriptional regulation of skeletal myogenesis. Curr. Opin. Genet. Dev. 2005, 15, 528–535. 10.1016/j.gde.2005.04.015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Rato L.; Duarte A. I.; Tomás G. D.; Santos M. S.; Moreira P. I.; Socorro S.; Cavaco J. E.; Alves M. G.; Oliveira P. F. Pre-diabetes alters testicular PGC1-α/SIRT3 axis modulating mitochondrial bioenergetics and oxidative stress. Biochim. Biophys. Acta 2014, 1837, 335–344. 10.1016/j.bbabio.2013.12.008. [DOI] [PubMed] [Google Scholar]
  57. Kim M.; Lim J.; Youn H.; Hong Y.; Yang K.; Park H.; Chung S.; Koh S.; Shin S.; Choi B.; et al. Resveratrol prevents renal lipotoxicity and inhibits mesangial cell glucotoxicity in a manner dependent on the AMPK–SIRT1–PGC1α axis in db/db mice. Diabetologia 2013, 56, 204–217. 10.1007/s00125-012-2747-2. [DOI] [PubMed] [Google Scholar]
  58. Blake R.; Trounce I. A. Mitochondrial dysfunction and complications associated with diabetes. Biochim. Biophys. Acta 2014, 1840, 1404–1412. 10.1016/j.bbagen.2013.11.007. [DOI] [PubMed] [Google Scholar]
  59. Li X.; Xu Z.; Jiang Z.; Sun L.; Ji J.; Miao J.; Zhang X.; Li X.; Huang S.; Wang T.; et al. Hypoglycemic effect of catalpol on high-fat diet/streptozotocin-induced diabetic mice by increasing skeletal muscle mitochondrial biogenesis. Acta Biochim. Biophys. Sin. 2014, 46, 738–748. 10.1093/abbs/gmu065. [DOI] [PubMed] [Google Scholar]
  60. Spiegelman B. M.; Heinrich R. Biological control through regulated transcriptional coactivators. Cell 2004, 119, 157–167. 10.1016/j.cell.2004.09.037. [DOI] [PubMed] [Google Scholar]
  61. Cantó C.; Auwerx J. PGC-1alpha, SIRT1 and AMPK, an energy sensing network that controls energy expenditure. Curr. Opin. Lipidol. 2009, 20, 98. 10.1097/MOL.0b013e328328d0a4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  62. Ma J.; Meng X.; Kang S. Y.; Zhang J.; Jung H. W.; Park Y.-K. Regulatory effects of the fruit extract of Lycium chinense and its active compound, betaine, on muscle differentiation and mitochondrial biogenesis in C2C12 cells. Biomed. Pharmacother. 2019, 118, 109297 10.1016/j.biopha.2019.109297. [DOI] [PubMed] [Google Scholar]
  63. Zhang J.-h.; Xin H.-l.; Xu Y.-m.; Shen Y.; He Y.-Q.; Lin B.; Song H.-t.; Yang H.-y.; Qin L.-p.; Zhang Q.-y.; et al. Morinda officinalis How.–A comprehensive review of traditional uses, phytochemistry and pharmacology. J. Ethnopharmacol. 2018, 213, 230–255. 10.1016/j.jep.2017.10.028. [DOI] [PubMed] [Google Scholar]
  64. Zhang H.-L.; Qing-Wen Z.; Xiao-Qi Z.; Wen-Cai Y.; Yi-Tao W. Chemical constituents from the roots of Morinda officinalis. Chin. J. Nat. Med. 2010, 8, 192–195. 10.3724/SP.J.1009.2010.00192. [DOI] [Google Scholar]
  65. Rusbana T. B.; Agista A. Z.; Saputra W. D.; Ohsaki Y.; Watanabe K.; Ardiansyah A.; Budijanto S.; Koseki T.; Aso H.; Komai M.; et al. Supplementation with fermented rice bran attenuates muscle atrophy in a diabetic rat model. Nutrients 2020, 12, 2409 10.3390/nu12082409. [DOI] [PMC free article] [PubMed] [Google Scholar]

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Supplementary Materials

ao1c03372_si_001.pdf (665.1KB, pdf)

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