Influence of mulberry leaf extract on serum adiponectin, visfatin and lipid profile levels in type 2 diabetic rats

Sepideh Barzin Tond Soudabeh Fallah Zahra Salemi Morteza Seifi About the authors

ABSTRACT

The effect of ethanolic mulberry leaf extract (MLE) and mulberry leaf powder (MLP) on glycemic control, serum adiponectin, visfatin and lipid profile in type2 diabetic rats have been investigated. 30 male wistar rats randomly divided into 5 groups. One group was randomly assigned as control (I) and diabetes was induced in others by administration of streptozotocin (STZ) (55 mg/kg body weight) 15 minutes after the administration of nicotinamide (110 mg/kg body weight) intraperitoneally. Finally, fasting blood glucose (FBG), lipid profile, adiponectin and visfatin were assessed after 6 weeks. Lipid profiles including serum FBG, total cholesterol, triglycerides, low-density lipoprotein cholesterol (LDL-c), very low-density lipoprotein cholesterol (VLDL) and visfatin significantly decreased and high-density lipoprotein cholesterol (HDL-c) and adiponectin increased in the two groups of treated diabetic rats in comparison to the diabetic control (p<0.05). For all the investigated factors, there was no significant difference between two treatment methods. However, MLP was more effective than MLE in improving visfatin. Results showed that MLE and MLP possess hypoglycemic and hypolipidemic activities and play an important role in regulating the secretion of adipokines such as adiponectin and visfatin.

Key words:
Mulberry leaf extract; mulberry leaf powder; type2 diabetes; adiponectin; visfatin

INTRODUCTION

Type 2 diabetes mellitus (T2DM) is one of the most common health problems in most countries of the world. Global prevalence of diabetes is predicted to increase to 366 million by 2030, largely owing to an aging population, increased urbanization and more sedentary lifestyles (11. Wild S, Roglic G, Green A, Sicree R, King H. Global Prevalence of Diabetes: Estimates for the year 2000 and projections for 2030. Diabetes Care 2004;27:1047-53.). 90-95% of diabetic cases suffer from T2DM, which is a heterogeneous disorder characterized by impaired cellular responses to insulin known as insulin resistance and followed by progressive partial pancreatic beta cell dysfunction (22. Chen D, Wang M-W. Development and application of rodent models for type 2 diabetes. Diabetes Obes Metab 2005;7:307-17., 33. Defronzo RA. Pathogenesis of type 2 diabetes mellitus. Medl Clin N Am 2004;88:787-835.). T2DM is mainly characterized by the development of increased morbidity and mortality for cardiovascular disease; it is also characterized by dramatic microangiopathic complications, such as retinopathy, nephropathy, and neuropathy (44. Chaturvedi N. The burden of diabetes and its complications: Trends and implications for intervention. Diabetes Res Clin Pr 2007;76:S3-S12.).

Evidence for the importance of plant extracts in the management of type 2 diabetes is emerging. Medicinal plants are frequently considered to be less toxic and free from side effects than synthetic ones (55. Iwu MM. Handbook of African medicinal plants. 2td ed. US: Boca Raton; 1993.). World health organization (WHO) encourages, recommends and promotes traditional / herbal remedies in natural health care programs because these drugs are easily available at low cost, safe and people have faith in them (66. Panchawat S RK, Sssisodia RK. Standardization and evaluation of herbal drug formulations. Alter Med 2010;15:43-7.).

Flavonoids are polyphenolic compounds with low molecular weight and play a major role in the cell synthesis (77. Fernández SP, Wasowski C, Loscalzo LM, Granger RE, Johnston GAR, Paladini AC, et al. Central nervous system depressant action of flavonoid glycosides. Eur J Pharmacol 2006;539:168-76.). These compounds are abundant in fruits and vegetables and increasing evidence demonstrates a positive relationship between consumption of flavonoids-rich foods and disease prevention (88. Babu PVA, Liu D, Gilbert ER. Recent advances in understanding the anti-diabetic actions of dietary flavonoids. J Nutr Biochem 2013;24:1777-89.). As for as diabetes mellitus is concern, flavonoids play a vital role in all aspects and its mechanism is well known (99. Patra JC CB. Artificial neural network-based drug design for diabetes mellitus using flavonoids. J Comput Chem 2011;32:555-67.). Recent reports indicate that mulberry leaves and leaf-dried extracts are a rich source of polyphenol antioxidants, including phenolic acids and flavonoids, such as Caffeic acid, 5- Caffeoylquinic acid , Kaempferol 3-O-(6"-malonyl-glucoside), Quercetin 3-O-(6″-O-malonyl)-β-D-glucoside (1010. Thabti I, Elfalleh W, Hannachi H, Ferchichi A, Campos MDG. Identification and quantification of phenolic acids and flavonol glycosides in Tunisian Morus species by HPLC-DAD and HPLC-MS. Functional Foods 2012;4:367-74.). Mulberry leaf, contains 18 sugar-mimic alkaloids and other active compounds, such as dietary fiber, rutin, isoquercitin, and astragalin. 1-deoxynojiricmycin (DNJ) is present in high concentration, accounting for 50% of iminosugars (1111. Asano N, Yamashita T, Yasuda K, Ikeda K, Kizu H, Kameda Y, et al. Polyhydroxylated Alkaloids Isolated from Mulberry Trees (Morus alba L.) and Silkworms (Bombyx mori L.). J Agr Food Chem 2001;49:4208-13.). The leaves are a valuable, yet low-cost material that can be used in reducing the risk and treatment of T2DM, and cardiovascular system, nervous system, as well as in weight loss (1212. Skowron MJ FE, Jeszka J, Krejpcio Z, Król E, Buchowski MS. Mulberry leaf extract intake reduces hyperglycaemia in streptozotocin (STZ)-induced diabetic rats fed high-fat diet. Functional Foods 2014;8:9-17.). Adipocytes produce and release a variety of proteins, collectively termed adipokines that exhibit important metabolic and inflammatory properties (1313. Lau DCW, Dhillon B, Yan H, Szmitko PE, Verma S. Adipokines: molecular links between obesity and atheroslcerosis. Am J Physiol - Heart 2005;288:H2031-H41.). Adiponectin is a 244 -residue protein and unlike most other adipokines, circulating adiponectin levels tend to be low in obese patients and increase with weight loss and with the use of insulin-sensitizing drugs. It increases fatty acid oxidation and glucose uptake in the muscle and reduces the synthesis of glucose in the liver (1414. Oh DK, Ciaraldi T, Henry RR. Adiponectin in health and disease. Diabetes Obes Metab 2007;9:282-9.). Visfatin is mainly produced by visceral fat, but it can be found in liver, skeletal muscle, bone marrow and lymphocyte. It exerts insulin-mimetic effects in vivo and in vitro and is upregulated in obesity, metabolic syndrome, and diabetes. Visfatin and insulin at similar concentrations have a comparable ability to induce glucose uptake and to inhibit glucose release. Visfatin activates insulin receptor though visfatin binds to this receptor at a distinct site from that of insulin. It is also implicated in dyslipidemia, hypertension, and generally atherosclerotic-related diseases. Subjects with metabolic abnormalities exhibit increased visfatin concentrations (1515. Filippatosm TD DC, Kiortsis DN, Tselepis AD, Elisaf MS. Increased plasma levels of visfatin/pre-B cell colony-enhancing factor in obese and overweight patients with metabolic syndrome. J Endocrinol Invest 2007;30:323-26.). We have recently shown that biochanin A possessed hypoglycemic and hypolipidemic activities and increased visfatin secretion (1616. Azizi R, Goodarzi MT, Salemi Z. Effect of Biochanin A on Serum Visfatin Level of Streptozocin-Induced Diabetic Rats. Iranian Red Crescent Me. 2014;16:e15424.). Treatment of diabetes has been under scientific investigation, and several strategies are being developed to manage it. Dietary modulation is the first -treatment option. Polyphenol derived from plants, have been put on test. In the present study, we aimed to investigate the effect of MLE and MLP on adipokine secretion in rats, with particular attention on the adiponectin and visfatin, each with unique properties.

MATERIAL AND METHODS

Chemicals and Reagents

Streptozotocin and nicotinamide were purchased from Sigma-Aldrich. All other chemicals used in this study were of analytical grade obtained from E Merck. Serum concentration of fasting blood glucose (FBG), total cholesterol, triglycerides (TG), and high-density lipoprotein cholesterol (HDL-c) were measured enzymatically using commercial kits (Pars Azemoon, Tehran, Iran) and spectrophotometer (JENWAY 6505, Europe Union). The serum low-density lipoprotein cholesterol (LDL-c) and very low-density lipoprotein cholesterol (VLDL) were calculated by the Friedewald formula (1717. Friedewald WT, Levy RI, Fredrickson DS. Estimation of the Concentration of Low-Density Lipoprotein Cholesterol in Plasma, Without Use of the Preparative Ultracentrifuge. Clin Chem 1972;18:499-502.) as follows:

LDL-c = Total cholesterol - [HDL-c + (TG/5)]

VLDL-c = TG/5

Insulin was determined by ELISA (Bioassay technology laboratory Shanghai, China). The concentration of serum adiponectin and visfatin was determined by commercial enzyme immunoassay kits (Bioassay technology laboratory Shanghai, China) and Elisa plate reader Bio Tek ELX800TM (VT, U.S.A).

Preparation of Mulberry Leaf Extracts (MLE) and Isolation

Leaves of Morus alba were collected from a farm in Tehran, Iran. It was washed thoroughly under running tap water, shade dried, and ground to a fine powder using an electric blender. 2200g of dried leaves powder were extracted three times with 96%ethanol by maceration at room temperature. The mixture was filtered, evaporated in vacuum evaporator to give 112g of extract. The obtained dry extract was suspended in water followed by extraction with hexane, CHCl3 and ethyl acetate for three times consecutively (18).

Measurement of Total Flavonoids Content

The total flavonoids content (TFC) in ethanolic extract of mulberry leaves and its fractions were determined using AlCl3 reagent. Briefly, 2.5 ml of each sample (and/or quercetin as the standard), previously dissolved in 90% ethanol, was mixed with 2.5 ml of a 2% AlCl3 solution in 90% ethanol. After 40 min, the absorbance of the yellow color was measured at 415 nm. The TFC [as μg quercetin equivalents/mg of sample] for the sample was calculated on the basis of a linear calibration curve obtained using quercetin (y=0.0169x+0.3526, r2=0.995) (19). The highest amounts of flavonoids were found in total extract and ethyl acetate extract respectively (Table 1). The extract was kept in 4C until further used.

Preparation of Mulberry Leaf Powder (MLP)

Dried mulberry leaf powder was administered by inclusion in the diet. To prepare MLP, standard feed was mixed with powdered mulberry leaves at 25% level (20).

Animals

Male Wistar rats (200-250 g) were purchased from the central animal house, Tehran University of Medical Sciences, and maintained in an air conditioned room (23±2ºC) and humidity (55±5%) with a 12 h light/12 h dark cycle. All the rats were provided with commercially available rat normal pellet diet (which contained carbohydrate 60% (w/w), fat 12% (w/w), protein 17.5% (w/w) and fiber 8% (w/w)) and water ad libitum. The study protocols were approved by the institutional animals' ethics committee of Arak University of Medical Sciences, Arak, Iran (With protection code of animal subject in medical research 93-176-32).

Induction and selection of type 2 diabetic rats

Type2 diabetes was induced in male wistar rats by a single-day intraperitoneal (IP) injection of

55mg/kg b.w. STZ, 15 minutes after the IP administration of 110 mg/kg b.w. of nicotinamide.

STZ was dissolved in citrate buffer, pH 4.5 and nicotinamide was dissolved in normal saline, while the respective control rats were given vehicle citrate buffer and normal saline (21). Diabetes induction was confirmed through measurement of blood glucose level with glucometer and rats with blood glucose levels more than 126 mg/dl were considered diabetic and selected for further studies (22).

Experimental design

In this experimental study, 30 male wistar rats randomly divided (simple randomization) into 5 groups (6 rats in each group). One of them was selected randomly as control (group1) and type 2 diabetes was induced in four other groups by administration of streptozotocin (55mg/kg of body weight) and nicotinamide (110 mg/kg body weight) intraperitoneally. The studied groups were: Group1. control (non-diabetic) (receiving normal diet and water ad libitum; Group 2. diabetic control (receiving normal diet and water ad libitum); Group 3. sham: diabetic (0.4 ml ethanol and 0.1 ml water); Group 4. diabetic + MLE (600mg /kg MLE dissolved in 0.4 ml ethanol and 0.1 ml water) and Group 5. diabetic + MLP (standard feed mixed with powdered mulberry leaves at 25% level). Treatments were continued for 6 weeks (Table 2). The initial and final body weight and FBG of all rats were determined before and after treatment, in fasting condition. Blood glucose level was determined using a glucometer (Accu-Chek Advantage II, Roche Diagnostics, Mannheim, Germany). The animals were anesthetized using Ketamine (75 mg/kg b.w) and Xylazine (10 mg/ Kg b.w) intraperitoneally. Blood sample was collected by cardiac puncture and serum was separated immediately.

Statistical analysis

All the data were expressed as mean± standard deviation (S.D.) of three replicates for six rats in each group. Stata software, version 13 (Stata Corp, College Station, TX, USA) was used for all statistical analysis. Normality assumption was checked using Shapiro Wilk test. One-way ANOVA was applied for determining differences between mean of the variable sin the studied groups. Post hoc Test (Tukey) was used to compare the data. Values of p<0.05 were considered statistically significant.

RESULTS

Fasting Blood Glucose Levels

Table 3 presents the effect of MLE and MLP on changes of fasting blood glucose in normal and diabetic rats. Serum glucose level was measured in all rats on 0 day and 42thday of oral administration of MLE and MLP. Three days after STZ injection, fasting blood glucose significantly increased in diabetic rats when compared to control group (p=0.001) (0day). Oral administration of MLE and MLP for 42 days, showed highly significant effect and FBG decreased in treated rats compared to untreated diabetic rats (P=0.001). No significant difference was observed between two treatment methods.

Serum Insulin Levels

Serum insulin levels decreased significantly in diabetic rats and after treatment with MLP and MLE for 42 days, insulin secretion increased significantly in all treated diabetic rats. Oral administration of MLP was more effective in insulin secretion than MLE, but the difference was not significant (data not shown).

Serum Lipids Levels

The effect of MLP and MLE administration on lipid profile was shown in table 4. The total cholesterol, triglyceride, VLDL and LDL concentrations in the serum were significantly higher in type-2 diabetic rats than in the control rats (P=0.001). The administration of MLE and MLP suppressed the increase in the total cholesterol, triglyceride, VLDL and LDL levels in the serum of diabetic rats. MLE and MLP administration decreased total cholesterol and triglyceride levels significantly to almost the control levels. Serum HDL-c was significantly lowered by diabetes induction (P=0.001); however, it was higher in MLE and MLP supplemented groups compared to the untreated diabetic groups (P=0.001). There was no significant difference between the effect of MLE and MLP administration in improving lipid profile.

Adiponectin and Visfatin Levels

Table 5 shows the effect of MLP and MLE on changes in adiponectin and visfatin concentration in normal and diabetic rats. Serum adiponectin levels were found to be reduced in diabetic rats significantly (P=0.001). The mean serum level of adiponectin in groups receiving MLE and MLP treatment were significantly increased, when compared to untreated diabetic rats (P=0.001), but there was no significant difference between MLE and MLP in increasing adiponectin levels in serum. The visfatin levels significantly increased in diabetic rats when compared to control group (P=0.001). Oral administration of MLP was significantly effective in 42 days and it returned to near normal in diabetic rats (P=0.001). MLE supplementation was also significantly effective in reducing visfatin levels (P= 0.001), but MLP was more effective than MLE (P= 0.002).

DISCUSSION

To determine the effect of various therapeutic agents such as MLE and MLP, we used nicotinamide and streptozotocin (STZ) to induce diabetes mellitus in rats. Streptozotocin-nicotinamide (STZ-NA)-induced diabetic rats exhibited moderate hyperglycemia associated with the less of postprandial early phase insulin secretion, as well as 50% decrease in pancreatic insulin content (2323. Oguri S, Motegi K, Endo Y. Augmented lipopolysaccharide-induction of the histamine-forming enzyme in streptozotocin-induced diabetic mice. BBA-Mol Basis Dis 2003;1637:83-90.). The results from this study suggest that MLE and MLP exhibit significant anti-diabetic properties by decreasing blood glucose levels, increasing the plasma insulin level and improving lipid profile in STZ-NA diabetic rats. Our results showed that, in all treated rats the increased levels of glucose were lowered significantly, but the level of FBG in treatment groups was still higher than the normal group. Additionally, in diabetic rats, degranulation or reduction of insulin secretion, arise from destruction of β-islet cells of pancreas. By the administration of MLE and MLP, the reduced glucose levels suggested that they likely increase the insulin secretion, which in turn, raise glucose uptake by tissues (2424. Lu M-P, Wang R, Song X, Chibbar R, Wang X, Wu L, et al. Dietary soy isoflavones increase insulin secretion and prevent the development of diabetic cataracts in streptozotocin-induced diabetic rats. Nutr Res 2008;28:464-71.). Elevation of the serum insulin in treated rats could be due to the insulinotropic substances, which induce the intact functional β-cells of the langerhans to produce insulin, or protect the functional β-cells from further deterioration, so that they remain active and produce insulin. It seems that the hypoglycemic effect of MLE and MLP are due to the increased level of serum insulin and the enhancement of peripheral metabolism of glucose (2525. Skim F LH, Kaaya A, el Amri H, Jana M. Pharmacological studies of two antidiabetic plants: Globularia alypum and Zygophyllum gaetulum. Therapie 1999;54:711-715.). On the other hand, the hypoglycemic activity of mulberry leaves maybe attributed to the high fiber content, the presence of trigonelline bases and moran A and/or moranoline in mulberry leaves. Mulberry leaf extract also contain other compounds with significant hypoglycemic activity in diabetic rats (2626. Mohammadi J, Naik P. Evaluation of hypoglycemic effect of &lt;i&gt; Morus alba&lt;/i&gt; in an animal model. Indian J Pharmacol 2008;40:15-8.). One of major constituent of Morus alba is deoxynojirimycin (DNJ) (2727. Butt MS, Nazir A, Sultan MT, Schroën K. Morus alba L. nature&apos;s functional tonic. Trends Food Sci Tech 2008;19:505-12.). DNJ, as a competitive inhibitor of intestinal α-glucosidase, affecting carbohydrate digestion and absorption, resulting in suppressed postprandial hyperglycemia. Intake of DNJ inhibited D-glucose uptake at the intestinal brush border membrane because of its similar size and, to some extent, structure to D-glucose (1212. Skowron MJ FE, Jeszka J, Krejpcio Z, Król E, Buchowski MS. Mulberry leaf extract intake reduces hyperglycaemia in streptozotocin (STZ)-induced diabetic rats fed high-fat diet. Functional Foods 2014;8:9-17.).

Diabetes is commonly associated with abnormalities in plasma lipid and lipoprotein levels. Abnormalities in lipid profile was associated with an increased risk of coronary heart disease, therefore ideal treatment for diabetes, in addition to glycemic control, should have a favorable effect on lipid profile (2828. Kesari AN, Kesari S, Singh SK, Gupta RK, Watal G. Studies on the glycemic and lipidemic effect of Murraya koenigii in experimental animals. J Ethnopharmacol 2007;112:305-11.). According to our results, diabetic condition in rats significantly increased serum total cholesterol (TC), LDL-c, TG and VLDL-C, while HDL cholesterol was significantly decreased. Administration of MLE or MLP improved these values to the normal ones. A significant reduction in serum triglyceride, total cholesterol, LDL-c and a significant increase in the HDL-c was observed. The triglyceride and cholesterol lowering effect of MLE and MLP is due to the stimulatory effect of quercetin on β-oxidation of fatty acid (1212. Skowron MJ FE, Jeszka J, Krejpcio Z, Król E, Buchowski MS. Mulberry leaf extract intake reduces hyperglycaemia in streptozotocin (STZ)-induced diabetic rats fed high-fat diet. Functional Foods 2014;8:9-17.). Several studies have already investigated various alkaloids, flavonoids and phytochemicals in white mulberry leaves to improve dyslipidemia, especially hypercholesterolemia (2929. Kobayashi Y, Miyazawa M, Kamei A, Abe K, Kojima T. Ameliorative Effects of Mulberry (Morus alba L.) Leaves on Hyperlipidemia in Rats Fed a High-Fat Diet: Induction of Fatty Acid Oxidation, Inhibition of Lipogenesis, and Suppression of Oxidative Stress. Biosci Biotech Bioch 2010;74:2385-95.) and inhibit oxidation of LDL-cholesterol (3030. Enkhmaa B, Shiwaku K, Katsube T, Kitajima K, Anuurad E, Yamasaki M, et al. Mulberry (Morus alba L.) Leaves and Their Major Flavonol Quercetin 3-(6-Malonylglucoside) Attenuate Atherosclerotic Lesion Development in LDL Receptor-Deficient Mice. J Nutr 2005;135:729-34.). The hypolipidemic effect of mulberry leaf extract in healthy non-diabetic human subjects was investigated and showed no significant difference in serum lipid profile, indicating the hypolipiemic effect of mulberry leaf tea may not be similar between normal and diabetic persons (3131. Kojima Y, Kimura T, Nakagawa K, Asai A, Hasumi K, Oikawa S, et al. Effects of Mulberry Leaf Extract Rich in 1-Deoxynojirimycin on Blood Lipid Profiles in Humans. J Clin Biochem Nutr 2010;47:155-61.). Rechel Dorothy et al. tested the hypolipidemic effects of mulberry leaf extract and found total cholesterol and triglycerides were significantly decreased in diabetic mulberry tea high group (DBTH), compared to diabetic control group (DBC) and diabetic mulberry tea low group (DMTL) and suggested the possible hypolipidemic effects of mulberry leaf tea where DNJ might be involved. Since it has been shown to be effective in decreasing lipid accumulation not only via increasing β-oxidation, but also by increasing adiponectin levels and activating AMP-activated protein kinase (AMPK) in isolated rat liver (3232. Tsuduki T, Nakamura Y, Honma T, Nakagawa K, Kimura T, Ikeda I, et al. Intake of 1-Deoxynojirimycin Suppresses Lipid Accumulation through Activation of the ß-Oxidation System in Rat Liver. J Agr Food Chem 2009;57:11024-9.). Insulin resistance maybe responsible for dyslipidemia, because insulin has an inhibitory action on HMG-coA reductase, a key enzyme in metabolism of cholesterol rich LDL-cholesterol particle (3333. Guoyan J. Practical diabetes mellitus. The People's Medical Publishing House, Beijing 1992.).

White adipose tissue is an active endocrine organ that secretes many biologically active mediators referred to as "adipokines" that have significant impact on energy consumption, carbohydrate and fat metabolism, glucose homeostasis and physiological functions such as immunity and inflammation (3434. Papanas N ME. Oral antidiabetic agents: anti-atherosclerotic properties beyond glucose lowering. Curr Pharm Design 2009;15:3179-92.). Adiponectin is defined as an anti-diabetic hormone secreted by adipose tissue. It was associated with many metabolic disorders, including obesity, insulin resistance, obesity related cardiovascular and fatty liver diseases (3535. Elissa LA, Elsherbiny NM, Magmomah AO. Propolis restored adiponectin level in type 2 diabetes through PPAR? activation. Egyptian Journal of Basic and Applied Sciences. 2015;2:318-26.). Our data showed that treatment with MLE and MLP, created a significant increase in serum level of adiponectin. Other studies also showed adiponectin has an adverse connection between FBG, TG, Total cholesterol and direct relation with HDL (3636. Eizadi M NF, Behboodi L , Khorshidi D. Correlation between serum adiponectin level and blood glucose concentration in adult asthmatic patients. Kashan University of Medical Sciences 2011;15:345-51.). Adiponectin concentration is known to be lower in insulin-resistant states such as obesity and T2DM (3737. Weyer C, Funahashi T, Tanaka S, Hotta K, Matsuzawa Y, Pratley RE, et al. Hypoadiponectinemia in Obesity and Type 2 Diabetes: Close Association with Insulin Resistance and Hyperinsulinemia. J Clin Endocr Metab 2001;86:1930-35.). A significant number of the metabolic actions of adiponectin are dependent on the activation of AMP-dependent kinase (AMPK) (3838. Yamauchi T, Kamon J, Minokoshi Y, Ito Y, Waki H, Uchida S, et al. Adiponectin stimulates glucose utilization and fatty-acid oxidation by activating AMP-activated protein kinase. Nat Med 2002;8:1288-95.). AMPK is a fuel-sensing enzyme and is activated by adiponectin, probably through a cAMP-dependent pathway (3939. Kahn BB, Alquier T, Carling D, Hardie DG. AMP-activated protein kinase: Ancient energy gauge provides clues to modern understanding of metabolism. Cell Metab 2005;1:15-25.). AMPK is activated when ATP is required, and one of its main stimulator is the AMP/ATP ratio, although it has been suggested that AMP kinase activation is the final common pathway of a number of insulin sensitizers including leptin and metformin (4040. Saha AK RN. Malonyl-CoA and AMP-activated protein kinase: an expanding partnership. Mol Cell Biochem. 2003;253:65-70.). Metformin increases glucose uptake, reduces hepatic glucose production and increases fatty acid oxidation with the aim of increasing ATP production. The reduction of hepatic and skeletal muscle triglyceride increases insulin sensitivity (4141. Yamauchi T, Kamon J, Waki H, Terauchi Y, Kubota N, Hara K, et al. The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity. Nat Med 2001;7:941-46.). Adiponectin also enhances the transcription of other genes involved in fatty acid metabolism, most notably peroxisome proliferator-activated receptor-α (PPARα) (4242. Yamauchi T, Kamon J, Ito Y, Tsuchida A, Yokomizo T, Kita S, et al. Cloning of adiponectin receptors that mediate antidiabetic metabolic effects. Nature 2003;423:762-69.), the target of the liberate group of lipid lowering agents. Activation of PPARα leads to an increase in levels of molecules involved in free fatty acid transport, such as CD36, and energy dissipation, such as uncoupling protein2 (4141. Yamauchi T, Kamon J, Waki H, Terauchi Y, Kubota N, Hara K, et al. The fat-derived hormone adiponectin reverses insulin resistance associated with both lipoatrophy and obesity. Nat Med 2001;7:941-46.) which also increases fatty acid oxidation. Adipocytes have an increasingly recognized role in the endocrine and paracrine control of metabolism and inflammation which appears to play a role in fatty acid and glucose metabolism through a change in insulin sensitivity and activation of fuel oxidation by AMPK and PPARα. Adiponectin has a plethora of anti-inflammatory and anti-atherogenic actions including inhibitors of tumor necrosis factor (TNF) signaling cascade, essentially acting as a TNF antagonist (4343. Gable DR, Hurel SJ, Humphries SE. Adiponectin and its gene variants as risk factors for insulin resistance, the metabolic syndrome and cardiovascular disease. Atherosclerosis 2006;188:231-44.). Our results showed treatment of diabetic rats with MLE and MLP significantly leads to elevated adiponectin level. There was no difference between two treatment methods. Effect of Morus alba leaves on adiponectin secretion in murine 3T3L1 cells showed they significantly stimulate lipid accumulation in cells and increase two key transcription factors involved in adipocyte differentiation, CCAAT/enhancer binding protein alpha (C/EBPα) and PPARγ, consequently. The expression of both aP2 and adiponectin is increased, as is the secretion of adiponectin. The results suggest that Morus alba leaves treatment may also improve insulin sensitivity by upregulating the secretion of adiponectin (44). Our results showed visfatin significantly increased in diabetic group and oral administration of MLE and MLP reduced it significantly, effectiveness of MLP was more than MLE in reducing visfatin. Studies revealed visfatin level increased in T2DM (45). Haider et al. showed that the release of visfatin by adipocytes in response to hyperglycemia is dependent on the duration and extent of glucose (4646. Haider DG SG, Kapiotis S, Maier C, Luger A, Wolzt M. The release of the adipocytokine visfatin is regulated by glucose and insulin. Diabetologia 2006;49:1909-14.). Some Other phenomena may also be involved in the increase of visfatin in type 2 diabetes. For instance, increase of visfatin levels may be a result of beta-cell deterioration, which is common in newly-diagnosed type 2 diabetic patients (4747. López-Bermejo A, Chico-Julià B, Fernàndez-Balsells M, Recasens M, Esteve E, Casamitjana R, et al. Serum Visfatin Increases With Progressive ß-Cell Deterioration. Diabetes. 2006;55:2871-5.). It has insulin mimetic effect and the affinity of visfatin for insulin receptor was found to be similar to that of insulin. Fukuhara et al. identified the potential role of visfatin as insulin mimetic and found that visfatin stimulated insulin receptor in a different way compared with insulin (4848. Fukuhara A MM, Nishizawa M, Segawa K, Tanaka M, Kishimoto K, Matsuki Y, et al. Visfatin: a protein secreted by visceral fat that mimics the effects of insulin. Science 2005;307:426-30.). Derdemezies et al. showed that quercetin reduces visfatin secretion. Similarly Chung et al. demonstrated that quercetin improves anti-inflammatory pathway and insulin resistance (4949. Derdemezis CS KD, Tsimihodimos V, Petraki MP, Vezyraki P, Elisaf MS, Tselepis AD. Effect of Plant Polyphenols on Adipokine Secretion from Human SGBS Adipocytes. Biochem Res Int 2011;2011.). In another study, Omer et al. showed quercetin shows protective effects in experimental diabetes, possibly by decreasing oxidative stress and preservation pancreatic β_cell integrity (50). We suggest that in the diabetic control group, due to increased blood glucose and decreased insulin levels, serum visfatin level was increased compensatory, whereas treatment with MLE and MLP led to decreased hyperglycemia and sensitivity and hence the expression of visfatin gene in treated groups decreased.

CONCLUSIONs

In Conclusion, our results showed that MLE and MLP possess hypoglycemic and hypolipidemic activities and play an important role in regulating the secretion of adipokines such as adiponectin and visfatin.

ACKNOWLEDGMENTS

The work was supported by grants from the Research Deputy of Arak University of Medical Sciences.

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Publication Dates

  • Publication in this collection
    2016

History

  • Received
    15 Jan 2016
  • Accepted
    25 Apr 2016
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