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CONCLUSION

REMISSION EFFECTS OF TRICHURIS SUIS (WHIPWORM) ON TYPE-1 DIABETES ON SPRAGUE-DAWLEY RATS


KIMBERLY S. JIMENEZ1 KEISHA MARIE G. TORRES1 and CRISTINA C. SALIBAY1 1Biological Sciences Department, College of Science, De La Salle UniversityDasmarias, Cavite

ABSTRACT
Remission effect of Trichuris suis ova on Type 1 diabetic Sprague-Dawley rats was determined in this study. Eighteen Sprague-Dawley rats obtained from the Bureau of Food and Drugs (BFAD) were induced for Type 1 Diabetes using Alloxan. During the three day post-induction, Trichuris suis ova was administered as treatment. Checking of blood glucose levels was done using a standard blood glucose meter once every week, for four weeks. The effects of Trichuris suis ova on Type 1 Diabetic Sprague-Dawley rats were monitored for four weeks. Results showed that the administration of 200 and 400 ova reduced blood glucose levels; however, the reduction was not sufficient to reduce blood glucose levels to normal. The treatment lost its efficacy during two week post-induction. The results revealed that 200 and 400 ova dosages were not sufficient to effectively treat diabetes since the glucose level reducing effects last only for 7-14 days.

Treatment group (no. of ova)

Glucose level 3 days Postinduction with Alloxan (mg/dl)

Number of Weeks Post-induction

Week 1 0 200 518a524.5a567.5a-

Week 2 563.5a-

Week 3 547.5a414.5a420.8a-

Week 4 740.5a400.17a460.4a-

200.17a+ 274.67a-

207.8b+ P<0.05 a320.0 *dissimilar400 letters (a,b) mean significantly different within treatment groups at a458.8
*dissimilar symbols (+, -) mean significantly different across treatment groups at P<0.05

INTRODUCTION
In the world, 246 million people are dealing with diabetes and 3.9 million of them reside in the Philippines. According to the Department of Health (DOH), 5 out of 100 deaths were caused by diabetes (Valisno 2010). People affected with Type 1 diabetes can inject insulin up to 1,500 times per year and prick their skin for blood 1,000 times (Lee 2008). People with Type 1 diabetes cannot produce their own insulin and therefore have to practice a rigid, daily regimen of exogenous insulin replacement and constant monitoring (UMMC 2009). Diabetes is a costly and time consuming affliction. Helminthic therapy, also called worm therapy, refers to the use of helminthes, which include hookworms and whipworms, in treating autoimmune related diseases such as mutliple sclerosis (MS), Crohns disease, asthma and allergies (Groce 2009). Cursory studies have been conducted regarding this field using Schistosomes, specifically, Schistosoma mansoni. Administration of Schistosome eggs in nonobese diabetic mouse models have been shown to prevent diabetes (Saunders et al. 2006). Unforutnately, Schistosomes are pathogenic and cause schistosomiasis in humans. The life cycle of Trichuris suis poses minimal risk of inadvertent colonization in humans, which makes them favorable for therapeutic use. Various studies have indicated that Trichuris suis ova (TSO) is a safe alternative for treatment of autoimmune diseases and immunological disorders such as such as Crohn's disease, multiple sclerosis, and asthma (Elliott et al. 2005). The primary aim of this study is to determine whether administering Trichuris suis ova is effective in inducing remission of type 1 diabetes in Sprague-Dawley rats.

Diabetes-induced rats treated with 400 ova showed a significant decrease (P<0.05) in glucose levels at week 1 by 251 mg/dl. The subsequent weeks (weeks 2 to 4) did not show any significant difference in glucose level compared to week 1, while a significant increase (P<0.05) by 252.6 mg/dl was observed at week 4. While glucose levels of diabetes-induced rats treated with 200 and 400 ova dosage showed a significant decrease of 367.33 g/dl and 359.7 g/dl respectively, against the control group ( P<0.05), the decrease in glucose levels between 200 and 400 ova dosages is not significantly different. The control group maintained its high blood glucose level beginning from post-induction to week 4. While the result shows no significant difference on the glucose levels after a longer period (2-4 weeks) post-induction, the increasing trend of glucose level is observed, which may be indicative of the lost of efficacy of the ova to reduce the glucose level as the time of administration of treatment become longer. Hence, regardless of dosage, there is a remarkable increasing trend in blood glucose levels as weeks progress. 800 700 600 500 Number of 400 Ova 300 200 100 0 Week 1 Week 2
Number of weeks post-induction

MATERIALS AND METHODS


Research Setting
The experimental procedure was performed at a well-ventilated improvised laboratory area in San Pedro, Laguna. 18 Sprague-Dawley rats were acquired from the Bureau of Food and Drugs (BFAD) Alabang, Muntinlupa City.

Research Procedure

Control 200 ova 400 ova Linear (Control) Linear (200 ova) Week 3 Week 4

Data Gathering and Statistical Analysis


The blood glucose levels of Sprague-Dawley rats were monitored using a device called Major II Blood Glucose Meter. Photodocumentation was performed using a digital camera. The paired t-test was used as a statistical analysis in determining the mean difference between the level of glucose before and after the administration of T. suis ova on the Sprague-Dawley rats. In determining the significant differences of the amount of T. suis ova administered on the rats in the level of glucose, the statistical tool used was one way ANOVA.

After induction of diabetes, the 200 ova treatment showed a slight decrease in glucose level during week 1, however, the change is not significant. In the subsequent weeks (weeks 2 to 4), the glucose levels increased and maintained hyperglycemia. Both the 200 and 400 ova treatment showed a lower glucose level when compared to the control during week one. Only treatment with 400 ova dosage induced lowering of the glucose level during week one. However, the effect is within a short period of time since during the succeeding weeks, there was an increasing trend of blood sugar level when no follow-up treatment was administered. Although, the 400 ova treatment has induced a decrease in glucose level, it is not sufficient to reduce the glucose level to normal (50-135 mg/dl). The result of the present study indicates that Trichuris suis ova was found to reduce the glucose level in Sprague-Dawley rats induced with Alloxan. However, the effect is within a short period of time since during the succeeding weeks, there is already an increasing trend of blood sugar level when no follow-up treatment was done.

Tail bitten by an aggressive rat

RESULTS and DISCUSSION


Pre- and Post-induction of Blood Glucose
All rats prior to treatment show normal glucose level ranging 66.17-94.17 mg/dl. The induction of type 1 diabetes significantly increased (P<0.05) the glucose level within treatment immediately after 3 days postinduction. An overall average increase of 431 mg/dl has been observed from the three groups. Expectedly, across treatment groups there is no significant difference (P>0.05) regarding the increase in glucose level. The data proves that alloxan has effectively induced diabetes. Treatment of Groups ( no. of Ova) Glucose level ** Glucose level (mg/dl) pre-induction 0 200 400 66.17a 94.17a 91.2a Glucose level (mg/dl) 3 days postinduction 518b 524.5b 458.8b significant;

CONCLUSION
Trichuris suis ova at 200 dosage reduces blood glucose level. However, the reduction is not statistically Trichuris suis ova at 400 dosage reduces blood glucose level however the reduction is not sufficient to
reduce blood glucose level to normal; Trichuris suis ova at 400 dosage loss its efficacy to reduce glucose level at week 2 post-induction

ACKNOWLEDGMENT
This work would not have been accomplished without the assistance of several people. We thank our thesis adviser Dr. Cristina Salibay, and our panel members Ms. Chona Bandelaria and Ms. Jonnacar San Sebastian. Dr. Cristina Salibay has been greatly responsible for the success of this study by providing us her expertise in research work. We express deep gratitude to our panel members for their patience and knowledge. Dr. Fedelino Malbas, a veterinarian, played an important role in this study by assisting us in inducing diabetes and administering treatment to the rats.

*Normal glucose level is 60-130 mg/dl ** dissimilar letters mean significantly different within treatment groups at P<0.05

BILIOGRAPHY
(1) Valisno J. 2010. Sweet danger. http://www.bworldonline.com/weekender/content. php?id=19511 (2) Lee MG. 2008. Why Diabetes scares even nondiabetics. http://library.pchrd .dost.gov.ph/index.php/newsarchive/851 (3) University of Maryland Medical Center. 2009. Diabetes-Type 1 Symtoms. http://www. Umm.edu/patiented/articles/what_symptoms_of_type_1_ diabetes_000009_4.htm (4) Groce V. 2009. Hygiene Hypothesis. Retrieved from http://foodallergies.about.com /od/foodallergybasics/f/hygienehypoth.htm (5) Saunders KA, Raine T, Cooke A, Lawrence CE. 2007. Inhibition of Autoimmune Type 1 Diabetes by Gastrointestinal Helminth Infection. Infection and Immunity 7 397-407. (6) Elliott DE, Summers RW, Weinstock JV. Dec 2006.Helminths as governors of immune-mediated inflammation. 2007 Australian Society for Parasitology Inc. Elsevier Ltd(whats the volume, page) this is a journal. 8p. (7) Ducommun D. 1992. Cage Hygiene, Healthy Litters, and Beddings. American Fancy Rat and Mouse Association Retrieved 12 February 2011 from http://www. afrma .org/rmindex.htm#health

Treatment with T. suis ova


After one week post-induction, rats receiving 200 ova dosage show decrease in blood sugar level. However, the decrease in blood glucose is not significant (P>0.05). During the subsequent weeks, the glucose levels of diabetic rats did not exhibit significant difference (P>0.05) relative to the first week.