Antidiabetic properties of an Ethanolic leaf extract of Launaea taraxacifolia (Willd.) Amin ex C. Jeffrey (Asteraceae) in SD rats

Diabetes mellitus (DM) is an autoimmune, hormonal, and metabolic disease [1]. There are two main types: Type 1 (insulin dependent) and Type 2 (non - insulin dependent), the most common form being the latter [2]. Type 2 diabetes, which affects 90–95% of diabetics, is characterized by inadequate glucose accessibility and utilization by target tissues due to β-cell dysfunction and insulin receptor resistance [3]. Poor management can result in retinopathies, nephropathies, neuropathies, cardiovascular challenges, and mortalities [4]. Non-conventional complications like dementia, hepatic abnormalities, cancers, and adverse geriatric conditions have recently been linked to poorly controlled diabetes [5]. Diabetes has become a public health problem globally. The WHO has warned that death due to diabetes in Africa will increase by 40% in the next ten years [3, 6]. However, the direct costs in case management in many developing countries is dominated by the relatively high costs of insulin and oral hypoglycaemic medicines at public health centres [7]. The populace rather depend on less expensive and readily available medicinal plant products for the management of diabetes and many other disease conditions which afflict them. Generally it is estimated that 70–80% of the world’s population use medicinal plants as the first line of treatment in their Primary Health Care system [6]. These medicinal plants are frequently considered to be less toxic and free from side effects than synthetic drugs [8‐12]. However, most of these medicinal plants used in the treatment of diseases in these developing countries have no scientific documented information. It is therefore imperative to investigate such medicinal plants in order to provide enough scientific data which may form the basis for the formulation of herbal hypoglycaemic products for clinical use after their safety has been ascertained.

African lettuce (Launaea taraxacifolia) is native to Africa, America, Europe, and Asia [13]. It is an annual herb that grows up to about 1 m tall and has a single glabrous stem that branches at the plant’s distal end. The leaves are linked to the aerial stem without a stalk. When studied microscopically, L. taraxacifolia is the only Asteraceae species without trichomes [14]. In many West African households, leaves of L. taraxacifolia are consumed raw in salads or cooked in stews and soups [15]. The leaves have also been used in traditional medicine to strengthen children’s bones [13] and to treat excessive blood pressure [16]. Adinortey and colleagues [12] revealed that L. taraxacifolia contains useful amounts of minerals and phytochemicals such as iron, magnesium, potassium, calcium, flavonoids, terpenoids, tannins, and saponins among others. It has been shown to possess antioxidant [17], anti-inflammatory [18], antimicrobial [18], anticancer [19], neuro-nephroprotective [20], and cardioprotective properties [21, 22]. A few investigators have reported on the blood glucose studies of L. taraxacifolia [16, 23, 24]. These studies generally used lower doses, and the reports are inconsistent. However, doses up to 5 g/kg are reportedly very safe and tolerable, with zero animal mortalities recorded in those studies [25, 26]. Due to the paucity and inconsistencies of such information on L. taraxacifolia, it was thus necessary to use higher doses to assess the antidiabetic properties of the leaf extract on glycemic control [27] and to further investigate the effects of the extract on pancreatic morphology [28], gluconeogenesis [29], intestinal glucose absorption [30], bodyweight [31] and dyslipidemia [32] that affect the status of diabetes. Therefore, the current study considered the potential antidiabetic activities of L. taraxacifolia and the possible mechanistic targets involved.

Type-2 diabetes continues to pose adverse microvascular and macrovascular challenges to patients, despite the availability of several orthodox antidiabetic medicines. Patients suffer from complications and adverse side effects, eventually requiring insulin shots, due to inadequate glycemic controls with the oral hypoglycemics [6, 7]. Plant-based therapies serve as alternatives but usually with a paucity of scientific evidence of efficacy. Given the scantiness and inconsistencies in the scientific evidence on the antidiabetic properties of L. taraxacifolia, the STZ-NAD model of type-2 diabetes [45] was used in this study to examine the effects of L. taraxacifolia extract on blood glucose. This study was also designed to further investigate the potential mechanistic targets of LTE in blood glucose regulation as well as its effects at the histological level.

Phytochemicals act as plant fingerprints [35] and offer lead compounds to develop conventional medicines [29]. The phytochemical analyses revealed the presence of alkaloids, saponins, glycosides, tannins, phytosterols, diterpenes, and flavonoids. These are secondary metabolites which are believed to be responsible for the observed effects in this study. The phytochemical results matched with previous research findings [12]. Other researchers have further identified compounds such as trans-9-tetradecenoic acid, cis-11-hexadecenoic acid, trans-2-octadecadecen-1-ol, oleic acid, palmitic acid, margaric acid, and stearic acid in the secondary metabolites of LTE using GC-MS analysis [16]. Catechin, caffeic acid, ellagic acid, quercetin and rutin have also been identified in LTE [46].

In the present study, the administration of STZ-NAD significantly increased plasma glucose levels sharply from 5 mmol/l to about 22 mmol/l. Streptozotocin, a known cytotoxic antibiotic with diabetogenic properties, is a glucose derivative that is selectively transported to pancreatic β-cells by GLUT-2 transporters [45]. In addition to alkylating and fragmenting DNA, STZ also generates reactive oxygen species (ROS) and nitric oxide and suppresses the activity of the enzyme aconitase [28]. On the other hand, Nicotinamide is a water-soluble B-vitamin with antioxidant and neuroprotective effects that offers partial protection to β-cells [28]. STZ-NAD is thus widely accepted as an experimental model of type-2 diabetes because it induces a near-60% decline in β-cell functions, accompanied by moderately high blood glucose levels [45].

A general reduction in blood glucose was noted following the administration of the different doses of LTE. The findings also revealed that LTE possesses anti-hyperglycemic potentials, as observed in the oral glucose tolerance test. These findings supported that of Gbadamosi et al. [47], who reported on the glucose-lowering and ameliorative potentials of some ethanolic fractions of LTE against diabetes complications. The authors attributed these effects to the extract’s ability to activate antioxidant enzymes. Again, using relatively lower doses of LTE in type-1 diabetic rats, Kuyoro et al. [24] noticed some anti-diabetic effects, but only after 2 weeks of treatment. However, Isehunwa et al. [23] and Koukoui et al. [16] found no antihyperglycemic effects with LTE treatment, using non-obese non-diabetic rats in a similar blood glucose study.

Anti-diabetic medications regulate blood glucose levels through various mechanisms [31]. The reduction in blood glucose levels seen in the rats given LTE might have happened via a single or multiple processes, including facilitation of insulin secretion, obstruction of intestinal glucose absorption, and inhibition of hepatic gluconeogenesis, among others. Several investigators have reported that certain plant medicines containing polyphenols improve insulin release [9, 11, 46]. As a result, the polyphenol phytochemicals in LTE, like glibenclamide (a long-acting sulfonylurea and an insulin secretagogue), may have worked on the residual β-cells to release insulin, facilitating glucose absorption [48]. Again, LTE and metformin were proven to lower fasting blood glucose levels. LTE phytoconstituents like flavonoids and tannins, as reported in other studies [9], may also have functioned similarly to metformin to lower glycemia by boosting peripheral glucose utilization [49].

Diabetes and cardiovascular diseases are strongly linked to dyslipidemia [27]. Other studies have found that type-2 diabetics have a high prevalence of abnormalities in plasma lipids such as total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterols (HDL-C), and low-density lipoprotein cholesterols (LDL-C) [50]. Alterations in serum lipids were noted following diabetes induction, which is consistent with earlier studies that diabetes promotes dyslipidemia [27]. The pathogenesis of diabetes has been strongly linked to the production of free fatty acids and low-grade inflammatory reactions as a result of dyslipidemia [51]. The improvements in lipid indices seen in the LTE-treated groups are comparable to what has previously been reported by Koukoui et al. [17].

Diabetes-related hepatic anomalies such as fatty liver disease, hepatic cirrhosis, and hepatocellular carcinomas [52] as well as STZ-induced liver damages [53] manifest as elevations in levels of liver enzymes such as AST, ALT, and ALP [52, 53]. These increments, as noted in this study, were attenuated with LTE administration. In addition, unlike the liver and kidney, the pancreas sizes shrank in the diabetic rats as has been found in certain people with type-2 diabetes [54]. Streptozotocin (STZ) is known to cause inflammatory reactions, lipid peroxidation, and severe antioxidant depletion in the Islet of Langerhans, which leads to apoptosis and necrosis [55, 56]. These findings are congruent with those shown in the photomicrographs of the pancreas from untreated diabetic rats, which showed severe pathological alterations in the Islet of Langerhans. LTE was able to moderate these developments. The observed beneficial qualities may be attributed to its antioxidant properties [17]. This is also consistent with recent findings that reducing oxidative stress enhances β-cell function regeneration [57].

Substantial weight loss was observed in all the diabetic rats. Even though weight loss was reversed in all treatment groups, the reversal was statistically significant only in diabetic rats fed with glibenclamide. This finding is consistent with previous studies, as glibenclamide has been linked to weight gain [48]. The poor reversal of weight loss noticed in diabetic rats treated with the extracts may also be a favorable feature of LTE because more than 80% of people with type-2 diabetes are already overweight or obese and require weight loss as part of their treatment plan [58]. Organ enlargements (hypertrophies) were noted in the livers and kidneys of the diabetic rats relative to their body weights. Hepatomegaly has been linked to about 70% of improperly treated type-2 diabetes cases [52]. Similarly, renal hypertrophy, which may be irreversible, is linked to diabetes [59]. The reduction in liver-to-body weight ratio and kidney-to-body weight ratio in the LTE treated groups suggests that the extract may be useful in the management of complications associated with diabetes, such as hepatomegaly and renal hypertrophy [52, 59].

Hepatic gluconeogenesis, which is increased in type-2 diabetes, tends to aggravate the disease [14]. Gluconeogenesis is mediated by enzymes such as pyruvate carboxylase, phosphoenolpyruvate (PEP) carboxykinase, fructose-1,6-bisphosphatase, and glucose-6-phosphatase [60]. The extracts decreased hepatic glucose synthesis, although in a concentration-independent manner. Phytochemicals like glycosides and triterpenes [9] in LTE, like metformin, may have activated the AMP Kinase, causing a switch from lipogenesis and gluconeogenesis to lipid oxidation and glucose absorption by muscle and liver cells [61]. In addition, LTE, like metformin, may have suppressed cAMP accumulation and hence lowered adenylate cyclase activity [9, 62].

While it is still unclear which of the phytoconstituents in this study were responsible for the anti-diabetic effects of LTE in the intestines, some investigators have reported on the ability of some flavonoids to inhibit the actions of glucose transporters, GLUT-2 and SGLT-1 [63, 64]. The LTE-mediated inhibition of glucose absorption by the whole intestines (in-vivo) and intestinal sacs (ex-vivo) may therefore have been similar to that of metformin, which is known to be deposited many folds into the brush borders of the intestines rather than plasma [65] and thus interferes with the actions of glucose transporters SGLT-1 and GLUT-2 to reduce blood glucose spikes [66]. Furthermore, Adinortey et al. [69] and Adedayo et al. [46] have also demonstrated that phytoconstituents in LTE inhibit gut enzymes including alpha-glucosidase, an enzyme found at the intestinal brush borders that plays a key role in converting complex sugars into glucose for easy absorption.

The similarities in results obtained using both the whole intestines and intestinal sacs suggest that the intestinal sac method may be a useful model for high throughput rapid screening of phytomedicines that may inhibit intestinal glucose absorption. Furthermore, the similarities suggest that the phytoconstituents in LTE may not be prodrugs, which require bioactivation to exert their pharmacological effects.

The extract lowered FBG levels and also suppressed hyperglycemia during the oral glucose tolerance test. LTE inhibited hepatic gluconeogenesis and suppressed intestinal glucose absorption. It also positively modulated levels of plasma lipids and liver enzymes. Although it did not improve body weight considerably, it reversed STZ-induced liver, kidney, and pancreatic damage and restored the morphology of the pancreatic Islet of Langerhans.

Future studies would consider a bioassay-guided fractionation to isolate and investigate the compounds responsible for the observed anti-diabetic and anti-lipidemic effects. The specific targets, enzymes and stages of gluconeogenesis and intestinal glucose transport interfered with by phytochemicals at the molecular level would also be investigated.