Effect of insulin sensitizer, rosiglitazone in streptozotocine induced diabetic db/db mice model

 

B. Jaynthi¹, P. Parameshwar², Tajuddin Baba M.D.¹, Kishore Pratap Patil¹, G. Cheranjeevi³

¹Annamalai University, Annamalai Nagar, Chidhambaram, Tamil Nadu, India

²Joythismathi Institute of Pharmaceutical Sciences, Timmapur, Karimnagar, A.P-505481

³Pulipati Prasad Institute of Pharmaceutical Sciences, Kammam

*Corresponding Author E-mail: param_pabba@yahoo.com

ABSTRACT:

Rosiglitazone is an oral drug that reduces the amount of sugar (glucose) in the blood. It is used for treating patients with type-2 diabetes and is in a class of anti-diabetic drugs called thiazolidinediones. Our diabetes clinic treatment algorithm mandates starting small doses of either a sulfonylurea agent or Metformin and increasing the dose every two weeks until either a fasting plasma glucose (FPG) concentration of <130 mg/dl is attained or a maximal (tolerated in the case of Metformin) dose is reached. The efficacy of thiazolidinediones, as compared with other oral glucose-lowering medications, in maintaining long-term glycemic control in type-2 diabetes is not known. We evaluated Rosiglitazone and metformin as initial treatment for type-2 diabetes mice. The potential risks and benefits, the profile of adverse events and the costs of these two drugs should all be considered to help inform the choice of pharmacotherapy for patients with type-2 diabetes. Rosiglitazone and Metformin are two pharmacologic agents useful in conditions characterized by insulin resistance. The aim of our study was to compare the efficacy of Rosiglitazone versus Metformin. Only the rosiglitazone group showed a significant reduction in insulin levels. Metformin and Rosiglitazone were well-tolerated.

 

 

INTRODUCTION:

Rosiglitazone works by helping to restore our body's proper response to insulin, thereby lowering our blood sugar. Effectively controlling high blood sugar helps prevent heart disease, strokes, kidney disease, blindness and circulation problems, as well as sexual function problems (impotence). Rosiglitazone is used either alone or in combination with other anti-diabetic medications (e.g., metformin, sulfonylureas, insulin)¹. Metformin is an oral diabetes medicine that helps control blood sugar levels. Metformin is for people with type 2 (non-insulin-dependent) diabetes. Metformin is sometimes used in combination with insulin or other medications, but it is not for treating type-1 diabetes².

 

Diabetes mellitus is a heterogeneous metabolic disorder characterized by altered carbohydrate, lipid and protein metabolism.

 

It is a common endocrine disorder and characterized by hyperglycemia resulting from defects in insulin secretion, insulin action or both. India has today become the diabetic capital of the world with over 20 million diabetics and this number is set to increase to 57 million by 2025. This astronomic increase in the prevalence of diabetes has made diabetes a major public health challenge for India³.

 

The Rosiglitazone exhibited 55.6 % glucose lowering in the db/db mice model as compared to the vehicle treated group because of the Rosiglitazone have the insulin sensitizer it increases the sensitivity of the insulin to the target tissues such as the liver, adipose tissue and muscle. Thiazolidinedione stimulate adipocyte differentiation, generating more numerous; smaller adipocyte. Insulin is more effective and there is less need for the β-cells increases the secretions of insulin and glucose level manages according to the insulin level⁴.

 

On the other hand metformin was exhibited 43.8 % of glucose lowering in the db/db mice  model due to it inhibit the hepatic gluconeogenesis that’s why glucose was not synthesized in the liver and it reduced the glucose level in the blood cells⁵.

 

The Rosiglitazone significantly exhibited an increase in the body weight as compared to vehicle treated group. It taken up free fatty acid and triglyceride and stored it in the adipocyte that’s why body weight gain occurred in Rosiglitazone treated group.

 

MATERIALS AND METHODS:

Preparation of test substances

Preparation of Rosiglitazone solution

The 10 mg/kg of rosiglitazone was dissolved in the 0.5 % carboxymethylcellullose solution using 1-2 drops of tween-80 as dissolution solvent. The dose of  Rosiglitazone was given the 5 ml/kg.

 

Preparation of the dose of metformin

The 500mg/kg of metformin was dissolved in 0.5 % carboxymethylcellullose solution using 1-2 drops of tween-80 as dissolution solvent. The dose of metformin was given the 5 ml/kg.

 

Induction of diabetes:

Diabetes was induced by a single i.p. injection of streptazocine at a dose of 60 mg/kg, freshly dissolved in citrate buffer (0.1M, PH 4.5). After 72 hours blood glucose was assayed and diabetes was verified.

 

Study design^6,7,8:

The study design was done for 07 days and daily body weight was recorded. The all treatments were administered two times a daily (9:00 am and 9:00 pm) through an oral route. The administration of vehicle and the entire drug was strated at 10 weeks of age. When diabetic state in db/db mice had been already established. For reference the control animals were administrated vehicle alone were also examined. Blood samples were collected in the fasting state from the retro orbital plexus by using heparinizd capillaries just before treatments. The blood samples were centrifuged at 4^oC on 8000 rpm for 7 minutes, and plasma was separated, frozen immediately, and stored at -70⁰c. These samples were analyzed for glucose, insulin, triglyceride, cholesterol, SGPT and SGOT. Plasma glucose and triglyceride levels were measured using a Hitachi 7350 auto analyzer. The insulin concentration was measured by Elisa kit.

 

Animals:

Female C57BL/6J db/db mice obtained from national institute of nutrition (Hyd India) were raised on standard laboratory chow. The animals were housed in cages in an air-conditioned room with a 12 hr light and dark cycle in specific pathogen free condition. The body weight of selected animal was in the range of 39-41 gm for experimental work.

Female db/db mice were divided into 3 groups,

Group I – Normal control healthy mice (8) received vehicle (0.5 % CMC)

Group II –Rosiglitazone treated mice (8), dose 5 mg/kg.

Group III-Metformin treated mice (8), dose 150 mg/kg.

 

Statistical analysis:

Data’s are presented as Mean ± SEM statistical analysis was performed with analysis of variance. Results obtained at the end of (day 7 and 10) experiments were compared with those obtained at zero days for same group and normal values, using student’s t-test for paired observations. 

 

RESULTS:

db/db mice model

Body weight         

The db/db mice were grouped on the basis of body weight. The body weight of the 3 groups of db/db mice at the beginning of the study did not differ significantly (36.6 = 0.3 g, 37.6 = 0.4 g and 36.0 = 0.2 g in the vehicle treated, Rosiglitazone-treated group showed significantly weight gain (p<0.05) than that of vehicle treated group. On the other hand Metformin did not show significant as compared to Rosiglitazone Table-1 and Fig. 1.

 

Plasma glucose: The change in plasma glucose level before to after treatment at day 0 to day 10 are shown in Table 2. Fasting plasma glucose level were significantly decreased in the Rosiglitazone group after treatment (p<0.001) as compare to vehicle group. Metformin also showed a significant reduction in plasma glucose level (p<0.005) as compare to vehicle treated group at day 10 (Table 2 and Fig. 2).

 

Triglyceride level

The triglyceride level in the Rosiglitazone group was significantly reduced than that in the vehicle treated group (p<0.001). Metformin also showed significant reduction in the triglyceride level (p<0.001) as compared with vehicle treated group (Table 3 andFig. 3).

 

Plasma Cholesterol

The Cholesterol level of Rosiglitazone treated group was significantly lowers (p<0.01) than the vehicle treated group at the treatment at day 10 and at day 5 (Table 4 and Fig. 4).

 

Plasma SGPT

The plasma SGPT level of Rosiglitazone treated group was significantly increased almost half after the treatment as compared to vehicle treated group. On the other hand Metformin did not show significant difference (Table 5 and Fig. 5).

 

Plasma SGOT level

The SGOT level was not significant at Day 10 as compared to Vehicle treated group, Rosiglitazone treated group and Metformin treated group. At the day 5 there was increase the level of SGOT in Rosiglitazone treated group as compare to vehicle treated group (Table 6 and Fig. 6).

 

Plasma insulin

The plasma insulin concentration in the Rosiglitazone treated group was significantly lower than that in the vehicle treated group and metformin group after treatment at the day 10 (p<0.01) (Table 7 and Fig. 7).

 

Table 1:  Mean body weight ± SEM

 

Table 2:  Mean plasma glucose ± SEM

 

Table 3:  Mean triglyceride levels ± SEM

 

Table 4: Mean cholesterol levels ± SEM

 

Table 5:  Mean plasma SGPT ± SEM

Groups	Treatment	Mean Plasma SGPT level at day 5 (mg/dl)	Mean Plasma SGPT  level at day 10 (mg/dl)
I II III	Vehicle Rosiglitazone Metformin	86.7 ± 11 109.8 ± 8.70 126.45 ± 46	105.45 ± 17.58 152.62 ± 17.89 93.9 ± 10.85

 

Table 6 Mean plasma SGOT ± SEM

Groups	Treatment	Mean Plasma SGOT  Level at day 5 (mg/dl )	Mean Plasma SGOT  level at day 10 (mg/dl)
I II III	Vehicle (I) Rosiglitazone (II) Metformin (III)	78.99 ± 6.80 110.25 ± 7.08 134 ± 37.33	98.71 ± 10.39 99.57 ± 5.97 93.95 ± 7.09

 

Table 7: Mean plasma insulin concentration at day 10 (ng/ml) ± SEM

 

Table 8:  Mean organ weight at day 10 ± SEM.

Groups	Treatment	Liver	Spleen	Heart	Kidney	White adipose tissue
I II III	Vehicle Rosiglitazone Metformin	2.1 ± 0.06 2.43 ± 0.14 2.40 ± 0.16	0.21 ± 0.10 0.11 ± 0.01 0.12 ± 0.02	0.12 ± 0.02 0.13 ± 0.04 0.12 ± 0.02	0.26 ± 0.02 0.25 ± 0.09 0.27 ± 0.08	1.26 ± 0.08 1.40 ± 0.18 1.10 ± 0.16

 

 

Fig. 1: db/db mice treated with Metformin and Rosiglitazine are in comparative with vehicle.

 

Fig. 2: Dissection view of treated db/db mice.

 

 

            

Fig.-1:  Mean body weight                                                                   Fig.-2:  Mean Plasma glucose

 

 

Fig.-3:  Mean triglyceride concentration                                               Fig.-4:  Mean cholesterol

 

Fig.-5:  Mean plasma SGPT                                                                       Fig.-6:  Mean plasma SGOT

 

 

Fig.-7:  Mean plasma insulin concentration                                            Fig.-8:  Mean organ weight

 

 

 

Organ weight

The liver weighty was increase in the Rosiglitazone treated group as compared to vehicle treated group. On the other hand Metformin did not show significant difference (Table 8 and Fig. 8)

 

DISCUSSION:

In the Rosiglitazone treated group there was a significant reduced in the concentration of insulin because of the db/db mice model was the insulin resistance there was a more amount of insulin is synthesized and the concentration of insulin was increased and it produces the resistance to the normal action of insulin in order Rosiglitazone acts as the insulin sensitizerand it reduced the level of insulin in db/db mice model. On the other hand Metformin was not more significant than Rosiglitazone treated group on the insulin concentration.

 

In the organ weight results there was no significant difference in the organ weight of liver, spleen, kidney, heartand white adipose tissue in the vehicle, Rosiglitazone and Metformin treated group. The SGPT results revealed that the Rosiglitazone was more significant than the Metformin. The Rosiglitazone was increases the concentration of serum glutamic pyruvate transferase (SGPT) in the treated group and that is efficacious for the diabetic patient. The SGOT results revealed that the Rosiglitazone was the only more significant than the Metformin andVehicle treated group at Day 5 only.

 

CONCLUSION:

The Rosiglitazone was significantly showed the decrease in the triglyceride level and cholesterol level in the db/db mice model. On the other hand Metformin significantly exhibited reduction in the level of the triglyceride but it did not affect the cholesterol level. Particularly in the vehicle treated group there was increased in the level of triglycerideand cholesterol at Day 10 in the bd/bd mice model.

 

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Received on 27.12.2011          Modified on 12.01.2012

Accepted on 12.02.2012         © RJPT All right reserved