Betaine treatment decreased serum glucose and lipid levels, hepatic and renal oxidative stress in streptozotocin-induced diabetic rats

Introduction

Diabetes mellitus (DM) is a chronic metabolic disease with high glucose levels caused by partial or complete insufficiency of insulin. Oxidative stress induced by hyperglycemia is accepted to play a major role in functional and structural defects seen in patients with DM and experimental models of diabetes [1], [2], [3]. Although exact mechanisms are unknown, autooxidation of glucose induced by hyperglycemia, formation of advanced glycation end products (AGEs), and activation of polyol pathway are considered to be responsible for the oxidative stress in DM [1], [2], [3].

Liver dysfunction and nephropathy are common complications of DM. Diabetic nephropathy is described with proteinuria, hypertension, and loss in renal function. It has been shown that DM diminishes prooxidant-antioxidant balance and causes inflammatory, apoptotic, and degenerative changes in liver [4], [5]. Similarly, DM was observed to increase oxidative stress and induce functional and structural defects in renal tissue [4], [6], [7], [8]. Based on these findings, several scientists have proposed antioxidant compounds to support treatment of DM. In this regard, effects of various antioxidant compounds (sulphydryl compounds, vitamin C and E, taurine, polyphenols) have been investigated [9].

Betaine (tri-methyl glycine; BET) is a choline metabolite formed in liver by the activities of mitochondrial choline dehydrogenase and betaine aldehyde dehydrogenase. Additionally, vegetables are alimentary sources of BET [10]. BET functions as an osmolite and shows antioxidant and antiinflammatory effects [10], [11]. Accordingly, BET treatment was reported to be useful against some oxidative stress-induced conditions such as atherosclerosis [12], liver [13], [14], [15], [16], and kidney [17], [18], [19], [20] disorders. In addition, it has been suggested that there is a relationship between the alteration of BET levels and development of DM complications [21], [22]. However, the effect of BET treatment on DM is poorly investigated [23], [24].

Metformin (MET) is an antidiabetic drug widely used in the treatment of DM [25]. MET was reported to ameliorate insulin resistance and have antioxidant, antiinflammatory and antiapoptotic effects [25], [26], [27]. These properties of MET contribute to its effect in preventing or ameliorating diabetic complications including hepatosteatosis [28], [29] and diabetic nephropathy [6], [28].

The present study aimed to investigate the ameliorative effects of BET on blood glycated hemoglobin (HbA_1c), serum glucose and lipid levels, oxidative stress, and hepatic and renal dysfunction in STZ-induced diabetic rats and the efficiency of BET was compared with MET, an oral antihyperglycemic drug with antioxidant potential.

Methods

Results

ROS, MDA, FRAP, and GSH levels in the liver and kidneys

Significant increases in ROS and MDA levels and decreases in FRAP and GSH levels were observed in the liver of STZ rats. Both BET and MET treatments decreased liver ROS and MDA levels, but GSH and FRAP levels remained unchanged (Figure 1). In kidneys of STZ rats, ROS and MDA levels also increased, but there were no changes in FRAP and GSH levels. Both BET and MET treatments decreased kidney ROS and MDA levels. Additionally, kidney GSH levels increased after MET treatment in STZ rats (Figure 2).

Figure 1:

The effects of metformin (MET) and betaine (BET) treatments on liver reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH), and ferric reducing antioxidant power (FRAP) levels in diabetic rats induced by streptozotocin (STZ) (Means±SE). ^ap<0.05 as compared to STZ group.

Figure 2:

The effects of metformin (MET) and betaine (BET) treatments on kidney reactive oxygen species (ROS), malondialdehyde (MDA), glutathione (GSH), and ferric reducing antioxidant power (FRAP) levels in diabetic rats induced by streptozotocin (STZ) (Means±SE). ^ap<0.05 as compared to STZ group.

Liver and kidney histology

In livers of STZ-diabetic rats, mild mononuclear inflammatory cell infiltration was observed in portal areas. Mild proliferation in biliary duct epithelium and mild inflammatory cell infiltration in portal areas were also seen. BET or MET treatment did not significantly affect these findings (Figure 3). Normal kidney histology was observed in STZ-diabetic rats. BET or MET treatments did not alter histologic findings in kidney (Figure 4).

Figure 3:

The effect of betaine (BET) and metformin treatments on liver histology in diabetic rats induced by streptozotocin (STZ) (H&E; ×100/×200).

Figure 4:

The effect of betaine (BET) and metformin treatments on kidney histology in diabetic rats induced by streptozotocin (STZ) (H&E; ×100/×200).

Discussion

STZ is a wide spectrum antibiotic frequently used to induce experimental DM. By damaging pancreatic β cells, STZ-induces a type 1-like DM. Thus, rat models of STZ-induced DM are convenient for studies on complications of DM [36]. A single STZ dose of 55–65 mg/kg is capable to induce functional and structural changes in different tissues of rats. In our study, rats were given 55 mg/kg STZ intraperitoneally. This treatment caused an increase in blood HbA_1c, and serum glucose and lipid levels, and a disturbance in examined liver and kidney function tests. Increases in ALT, AST, and LDH activities together with hepatic histopathologic changes indicated the presence of hepatic dysfunction in STZ rats. On the other hand, elevated serum BUN levels, diminished total protein and albumin levels, and increased urinary protein levels were observed despite normal histologic appearance of kidney sections and unchanged creatinine levels. As is known, elevated BUN and creatinine levels are markers for renal injury. Elevated creatinine levels are seen only in advanced stage kidney damage whereas BUN levels increase earlier. Therefore, elevated BUN levels together with normal kidney histopathology and unchanged creatinine levels, as seen in our study, has been suggested to indicate a mild renal injury [37]. Under these conditions, significant increases in ROS production and lipid peroxide levels were detected in the liver and kidney of DM rats, indicating a prooxidant status in these tissues. These findings are in accordance with previous studies [4], [5], [6], [7], [8].

In the current study, the effect of BET treatment on blood glycemia, lipids, hepatic and renal function tests as well as oxidative stress in liver and kidney tissues were evaluated in STZ-induced diabetic rats. Additionally, the efficiency of BET was compared to that of MET. It has been reported that MET ameliorated insulin resistance and had antioxidant, antiinflammatory and antiapoptotic effects [25], [38]. These properties of MET may contribute to its preventive or ameliorative effect on complications of DM including hepatic [5], [28] and renal complications [6], [39]. In the current study, MET ameliorated hepatic function tests and decreased high levels of serum glucose and triglyceride, blood HbA_1c, and urinary protein in STZ rats. Additionally, ROS and lipid peroxide levels diminished in the liver and kidney of MET-treated STZ rats. However, this treatment did not alter histopathologic appearance in these tissues.

On the other hand, BET has antioxidant and antiinflammatory effects. Although it does not interact directly with oxidants, the effect of BET on the metabolism of sulfur-containing substances in the liver was suggested to be responsible for its antioxidant activity [40]. BET administration decreases homocysteine levels, and increases S-adenosyl methionine (SAM) and GSH levels which have strong antioxidant effects in organism [10], [11]. Among them, SAM has a direct antioxidant activity, scavenges ROS and chelates iron ions and thereby inhibits hydroxyl radical generation [41]. Accordingly, BET was reported to protect tissues including liver and kidney against oxidative stress mediated injury. BET treatment was reported to be useful in the alleviation of alcoholic-[13] and non-alcoholic fatty liver [14], and necrotic [15] and fibrotic [16] lesions in the liver. Previous studies have also demonstrated protective effects of BET against carbon tetrachloride-[17], high fructose diet-[19] and cisplatin-[20]-induced nephrotoxicity.

Decreased liver [21] and kidney [42] levels of BET were reported in animal models of DM. In human studies, increased urinary excretion and decreased serum levels of BET were shown to be correlated with DM or its complications [22], [43], [44]. However, effects of BET supplementation in rats with DM have not been clearly elucidated yet. Jeong et al. [23] have found mildly decreased serum glucose and fructosamine levels, AST, ALT, and LDH activities together with amelioration in histopathology of pancreatic β cells in DM rats fed on 1% BET containing diet for 3 weeks. In another study, Jung et al. [24] have observed that 1% BET supplemented diet for 5 weeks resulted in decreases in insulin and triglyceride levels and amelioration of insulin resistance in db/db mice. They also found decreased lipid peroxide and increased GSH levels and catalase activities in the liver. However, BET treatment (500 mg/kg for 2 weeks with oral route) did not decrease blood glucose levels in STZ rats, but it prevented and/or delayed complications of diabetic retinopathy by inhibiting retinal neovascularization [45].

In the current study, BET treatment decreased blood HbA_1c, serum glucose and lipid levels, and urine protein excretion and reduced the prooxidant status in liver and kidney tissues of DM rats. However, it did not ameliorate hepatic and renal function tests in serum and histopathologic findings in the liver or kidney. As a well known antidiabetic drug, MET was used in our study to compare the effects of BET. Our results are showing that BET exerts effects similar to that of MET in STZ-induced diabetic rats, but they do not imply any superiority. Therefore, investigating the effects of the co-treatment of BET and MET in STZ-treated rats can be interesting.

In conclusion, all of the above mentioned findings indicate that BET may be a candidate supplement in the treatment of DM and prevention of its complications.