Elsevier

Journal of Ethnopharmacology

Volume 231, 1 March 2019, Pages 29-38
Journal of Ethnopharmacology

The methanolic extract of Thymus praecox subsp. skorpilii var. skorpilii restores glucose homeostasis, ameliorates insulin resistance and improves pancreatic β-cell function on streptozotocin/nicotinamide-induced type 2 diabetic rats

https://doi.org/10.1016/j.jep.2018.10.028Get rights and content

Abstract

Ethnopharmacological relevance

Thymus praecox subsp. skorpilii var. skorpilii (syn. Thymus praecox subsp. jankae (Celak.) Jalas) is consumed as a Turkish folk medicine for the treatment of spasm, sore throat and shortness of breath, also having strong antioxidant activity and the leaves of the plant have been utilized for the treatment of diabetes as the decoction in Turkey.

Aim of the study

In the present study, we aimed to investigate the potential mechanism of antidiabetic action of Thymus praecox subsp. skorpilii var. skorpilii methanolic extract (TPSE) on streptozotocin (STZ)/nicotinamide (NA)-induced type 2 diabetic rats.

Materials and methods

Sprague Dawley rats were randomly divided into four groups; control, diabetes, TPSE (100 mg/kg b.w, p.o.) and metformin group (400 mg/kg b.w, p.o.). Diabetes was established in all groups except control group by 55 mg/kg STZ (i.p.) for once 15 min after 100 mg/kg NA injection. 3 days after STZ/NA injection, treatments were administered for three weeks and then rats were decapitated; tissue and blood samples were obtained for measuring the level of glucose transporters (both GLUTs and sodium glucose co-transporters (SGLTs)), enzymes related to glucose (Hexokinase (HK), phosphoenolpyruvate carboxykinase (PEPCK), α-glucosidase) and lipid metabolism (Acetyl-coenzyme carboxylase (ACC)), AST, ALT, creatinine, insulin, anti-inflammatory (IL-10) and inflammatory (TNF-α, IL-1β, IL-6) cytokines, AMP-activated protein kinase (AMPK), peroxisome proliferator-activated receptor gamma (PPAR-γ) and glucagon like peptide-1 (GLP-1). Histopathological alterations of the pancreas were examined.

Results

After three weeks of treatment, TPSE has exhibited a significant reduction of plasma levels of the proinflammatory cytokines. Besides, TPSE treatment elevated plasma insulin levels and normalized blood glucose levels. Moreover, it improved the values of AMPK in liver and GLP-1 in pancreas. Increased α-glucosidase, PEPCK, GLUT-2 and SGLTs levels with the induction of diabetes considerably lowered with TPSE treatment. Especially on SGLT-2, TPSE achieved a more prominent decrease. After the atrophy in Langerhans islets due to diabetes induction, treatment was found to prevent the damage of islets.

Conclusions

Based on the findings presented here, it has been concluded that TPSE has marked antidiabetic effects through various pathways on STZ/NA-induced diabetic rats and it may potentially be used as an effective treatment for type 2 diabetes mellitus (T2DM). Further research on isolation of the bioactive components is underway.

Introduction

Diabetes mellitus (DM) is a metabolic disorder which occurs due to hyperglycemia resulting from insulin resistance and/or deficiency in insulin secretion (Alam et al., 2014). Several pathologic mechanisms have been implicated in diabetes. Dysregulation of hepatic glucose metabolism is a core defect in the development of insulin resistance and subsequently Type 2 Diabetes Mellitus (T2DM) (Petersen et al., 2017).

The rate limiting enzymes of several processes implicated in glucose metabolism become important in the view of T2DM. For instance, phosphoenolpyruvate carboxykinase (PEPCK), hexokinase (HK) and glycogen synthase are responsible for gluconeogenesis, glycolysis and glycogenesis, respectively (Bouskila et al., 2010, Kwon et al., 2017, Liu et al., 2014). The transport systems mediating glucose intake and utilization are classified into two groups: glucose transporter proteins (GLUTs) and sodium-dependent glucose transporter proteins (SGLTs) (Harada and Inagaki, 2012). According to the literature data, upregulation of these two transporters can cause hyperglycemia (Hansen et al., 2014, Peng et al., 2015).

The pharmacological agents, used currently for the treatment of T2DM include mainly oral anti-diabetic drugs, such as sulfonylureas, biguanides, thiazolidinediones, α-glucosidase inhibitors, and dipeptidyl peptidase-4 (DPP-4) inhibitors, have restricted usage due to their inefficacy at the progression of diabetic complications and undesired side effects including nausea, diarrhea, hypoglycemia, weight gain, liver and heart failure (Alhadramy, 2016, Chaudhury et al., 2017, Du et al., 2014).

In order to keep blood glucose levels under control in DM, about 1200 plants are used worldwide and approximately 30% of conventional antidiabetic plants are examined pharmacologically and chemically. Plants are attractive sources for the development of novel safe and effective medicines for DM owing to their effectiveness, minimal side effects in clinical experiences and also relatively low cost (Nasri et al., 2015).

The genus Thymus (Lamiaceae) comprises of about 928 species worldwide (Morales, 2002). The several major components in essential oil of the different Thymus spp. are thymol, carvacrol, linalool, geraniol, thujanol, p-cymene which are behind the therapeutic activities (Schmidt et al., 2012). They are commonly used as culinary herbs and flavouring agents. Due to their antimicrobial, spasmolytic and antioxidant effects, they have been used for medicinal purposes. Besides, anti-inflammatory, antinociceptive and antitumoral activities of Thymus spp. that may vary depending on the species and the amount of chemical compositions have been reported (Safaei-Ghomi et al., 2009).

As having different biological activities, Thymus spp. have been tested in diabetic conditions (Al-Mustafa and Al-Thunibat, 2008, Alamgeer et al., 2016, Hajializadeh et al., 2014a, Ozkol et al., 2013). Although antidiabetic activities of Thymus spp. are attributed to their antioxidant properties, the exact mechanisms have not been fully elucidated by now.

Thymus praecox subsp. skorpilii var. skorpilii (TPS) (syn. Thymus praecox subsp. jankae (Celak.) Jalas) is a plant with long±woody, creeping branches, non-flowering or with a terminal inflorescence, flowering stems are born in rows. Cauline leaves of plant lanceolate-obovate to ovate-elliptic or almost orbicular. Corolla mauve to purple, rarely whitish (Jalas, 1982).

The dried leaves of T. praecox subsp. skorpilii var. skorpilii is used in tea form for the purpose of antispasmolytic, softening the throat and shortness of breath in Eskişehir-Turkey in traditional medicine (Yucel et al., 2010). Besides, leaves and aerial parts of this plant are used for the treatment of diabetes in Amasya and Giresun provinces of Turkey (Ezer and Mumcu Arisan, 2006, Tuzlaci, 2016). Methanol and water extracts of the leaves of TPS has been reported to possess significant antioxidant effect, native to Thymus spp. In details, it has been found that water and methanol extracts of TPS possessed reducing power effect in a dose-dependent manner. In addition, the major components of essential oil of this species were identified as thymol, o-cymene, γ-terpinene and α-pinene using GC-MS technique (Ozen et al., 2011).

In our previous investigation, the phenolic characterization of the methanol extract from plant was performed by ESI-Q-TOF LC/MS technique. The phenolic compounds in methanol extract were identified as chlorogenic acid, luteolin-7-O- glucoside, 3-O-feruloylquinic acid, quercetin-3-O-hexoside, and apigenin-7-O-glucuronide (Ucman et al., 2017).

According to the recorded data, there is no study showing the mechanism of antidiabetic action of TPS. Shortly after, in our study, we firstly evaluated the antidiabetic effects of TPS in diabetic rats induced by streptozotocin (STZ)/nicotinamide (NA) and we clearly observed that TPS extract has remarkably decreased blood glucose levels (Cam et al., 2017).

By this way, in the present study, our aim was to uncover the mechanisms of action of TPS by monitoring the biochemical and histopathological parameters in diabetic condition induced by STZ/NA in rats.

Section snippets

Chemicals

STZ, NA, metformin, Gill's Hematoxylin Solution were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). Enzyme-linked immunosorbent assay (ELISA) kits were procured from Bioassay Technology Laborator, Korain Biotech Co., Ltd (Shanghai, China).

Plant material and extract preparation

The aerial parts of TPS were collected in 15 July 2016 from Uludag-Bursa, Turkey in blooming semester and identified by Assist. Prof. Dr. İsmail Şenkardeş, Marmara University, Faculty of Pharmacy, Pharmaceutical Botany Department. A voucher

Effects of TPSE on blood glucose levels

The rats were bled by orbital puncture under ether anesthesia for measuring blood glucose levels. Diabetic rats showed a significant increase (438.83 ± 17.83 mg/dL, p < 0.001) in blood glucose levels compared with the other groups at the end of three weeks (Table 1). Blood glucose levels of TPSE treated rats lowered considerably (205.67 ± 11.56 mg/dL) compared to diabetic rats after one week and these decreases continued for three weeks (132.17 ± 4.58 mg/dL). At the end of three weeks,

Discussion

Classical therapy for T2DM has some limitation and because of that there is a need to go beyond the current standard of care. In this regard, approximately 30% of diabetic patients utilize the complementary and alternative medicine (Shewamene et al., 2015). Therefore, many herbal extracts have been investigated on experimental models in the context of their anti-diabetic activity (Pang et al., 2015).

STZ/NA-induced rat model of T2DM is described one of the most appropriate experimental models to

Conclusion

As noted in this study, the antidiabetic effects of the methanolic extract of TPSE can restore glucose homeostasis, ameliorate insulin resistance and decrease pancreatic β-cell damage. These effects could be due to SGLT-1, SGLT-2, GLUT2, PEPCK, α-glucosidase inhibition and GLP-1 activation in STZ/NA-induced diabetic rats. In the light of these findings, we suggest that TPSE is beneficial in diabetes treatment and the effects may come from compounds of chlorogenic acid, luteolin-7-O- glucoside, 3

Acknowledgements

This study was supported by Marmara University, Scientific Research Projects Committee (MU-BAPKO; SAG-C-DRP-250416-0173).

Conflicts of interest

Authors all declare that there is no existing conflict of interest.

Author Contribution

Muhammet Emin Cam: 5 Ayse Nur Hazar-Yavuz: 13 Sila Yildiz: 14 Busra Ertas: 7 Betul Ayaz Adakul: 12 Turgut Taskin: 8 Levent Kabasakal: 10

References (67)

  • Y. Okamoto et al.

    Enhanced GLUT2 gene expression in an oleic acid-induced in vitro fatty liver model

    Hepatol. Res.

    (2002)
  • T. Ozen et al.

    Determination of antioxidant activities of various extracts and essential oil compositions of Thymus praecox subsp. skorpilii var. skorpilii

    Food Chem.

    (2011)
  • B.J. Peng et al.

    Chlorogenic acid maintains glucose homeostasis through modulating the expression of SGLT-1, GLUT-2, and PLG in different intestinal segments of Sprague-Dawley rats fed a high-fat diet

    Biomed. Environ. Sci.

    (2015)
  • S.K. Ramaiah

    A toxicologist guide to the diagnostic interpretation of hepatic biochemical parameters

    Food Chem. Toxicol.

    (2007)
  • J. Safaei-Ghomi et al.

    GC/MS analysis and in vitro antioxidant activity of essential oil and methanol extracts of Thymus caramanicus Jalas and its main constituent carvacrol

    Food Chem.

    (2009)
  • M. Snel et al.

    Immediate and long-term effects of addition of exercise to a 16-week very low calorie diet on low-grade inflammation in obese, insulin-dependent type 2 diabetic patients

    Food Chem. Toxicol.

    (2011)
  • N.M. Tabatabai et al.

    Enhanced expressions of sodium-glucose cotransporters in the kidneys of diabetic Zucker rats

    Diabetes Res. Clin. Pract.

    (2009)
  • A.H. Al-Mustafa et al.

    Antioxidant activity of some Jordanian medicinal plants used traditionally for treatment of diabetes

    Pak. J. Biol. Sci.

    (2008)
  • Alamgeer et al.

    Comparative hypoglycemic activity of different fractions of thymus serpyllum L. in alloxan induced diabetic rabbits

    Pak. J. Pharm. Sci.

    (2016)
  • R. Burcelin et al.

    Glucose sensing by the hepatoportal sensor is GLUT2-dependent: in vivo analysis in GLUT2-null mice

    Diabetes

    (2000)
  • M.E. Cam et al.

    Antidiabetic effects of Salvia triloba and thymus praecox subsp. skorpilii var. skorpilii in a rat model of streptozotocin/nicotinamide-induced diabetes

    Marmara Pharm. J.

    (2017)
  • A. Chaudhury et al.

    Clinical review of Antidiabetic drugs: implications for type 2 diabetes mellitus management

    Front. Endocrinol.

    (2017)
  • S.C. Cramer et al.

    Colocalization of GLUT2 glucose transporter, sodium/glucose cotransporter, and gamma-glutamyl transpeptidase in rat kidney with double-peroxidase immunocytochemistry

    Diabetes

    (1992)
  • C. Culling

    Handbook of Histopathological and Histochemical Techniques

    (1974)
  • M.C. Deeds et al.

    Single dose streptozotocin-induced diabetes: considerations for study design in islet transplantation models

    Lab. Anim.

    (2011)
  • Y.-F. Du et al.

    Achieving glycemic control in elderly patients with type 2 diabetes: a critical comparison of current options

    Clin. Interv. Aging

    (2014)
  • J. Dyer et al.

    Expression of monosaccharide transporters in intestine of diabetic humans

    Am. J. Physiol. Gastrointest. Liver Physiol.

    (2002)
  • N. Ezer et al.

    Folk medicines in Merzifon (Amasya, Turkey)

    Turk. J. Bot.

    (2006)
  • M. Garcia et al.

    Hypothalamic ependymal-glial cells express the glucose transporter GLUT2, a protein involved in glucose sensing

    J. Neurochem.

    (2003)
  • J.E. Gerich

    Role of the kidney in normal glucose homeostasis and in the hyperglycaemia of diabetes mellitus: therapeutic implications

    Diabet. Med.

    (2010)
  • V. Gorboulev et al.

    Na(+)-D-glucose cotransporter SGLT1 is pivotal for intestinal glucose absorption and glucose-dependent incretin secretion

    Diabetes

    (2012)
  • H.H. Hansen et al.

    The sodium glucose cotransporter type 2 inhibitor empagliflozin preserves beta-cell mass and restores glucose homeostasis in the male zucker diabetic fatty rat

    J. Pharmacol. Exp. Ther.

    (2014)
  • N. Harada et al.

    Role of sodium-glucose transporters in glucose uptake of the intestine and kidney

    J. Diabetes Investig.

    (2012)
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