Diabetes: Encapsulation to Treat a Disease

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Table of Contents

Introduction
Background and significance
Specific Aims
References

Introduction

The science of treating disorders has transformed into a rapid problem solving platform with the intervention of Biomedical engineering. This has initiated confidence in the health care providers and patients with the hope that they could overcome the complications of debilitating disorders. The pathological involvement of vital organs has prompted the researchers to investigate on the strategies that aim at replacing or repairing the tissues with functionally advanced counterparts. This would require an understanding of cellular structural frame work.

However, the approach of drug delivery or any agent intended for alleviating the health complications may also need to be considered in addition to tissue engineering.

For this purpose, there was a growing emphasis on the approach of encapsulation to treat a disease. The present description was focused on the utility and validity of encapsulation concerning Diabetes mellitus. This metabolic disorder has emerged as a major health burden in many countries due to its growing indices of prevalence. The function of key hormone, insulin gets altered leading to insulin resistance and abnormal glucose levels. This disorder may have etiologies of genetic, biochemical or immunological origin. This disorder was also believed to be associated with hypertension and cardiovascular defects. Although conventional therapies have contributed to the understanding and management of diabetes, there is also a need to emphasize on the treatment modalities keeping in view of encapsulation.

This could be because this encapsulation technique was believed to be primarily cost efficient and may serve as better strategy to address the issues concerned with the cure of diabetes. Therefore, the objective of this description was to propose a study for treating diabetes with special emphasis on islet cell encapsulation and the related aspects of tissue engineering.

Background and significance

Diabetes mellitus, considered as a disorder of glucose homeostasis was reported to result from the immune-mediated damage of pancreatic beta cells in the islets of Langerhans (type 1 diabetes) or the insulin resistance and obesity syndromes (Giannoukakis & Robbins, 2002designtheyalginate-based The involvement of the pancreas in the etiopathogenesis of diabetes has increased the focus on Islet cells. Hence, transplantation of islet cells to inhibit the immune-mediated graft rejection was considered a better option previously to restore the specific pancreatic function (Giannoukakis & Robbins, 2002).

Further, diabetes mellitus was associated with the risk of coronary heart disease (CHD) which was revealed by the elevated levels of serum C-reactive protein (CRP) (Bahceci et al., 2008). The utility of CRP was found to play a vital role in disorders where inflammation was reported. Hence, it was better regarded as an important inflammatory marker (Bahceci et al., 2008).

Blaha et al. (2008) reported a comprehensive management plan known as the “ABCDE” approach that could better assist physicians to gain insights on the metabolic syndrome that is associated with cardiovascular disease and type 2 diabetes mellitus. Here”A” stands for the assessment of cardiovascular risk and aspirin therapy, “B” for blood pressure control, “C” for cholesterol management, “D” for diabetes prevention and diet therapy, and “E” for exercise therapy.

These approaches may serve the best to provide a satisfactory remedy (Blaha et al., 2008).

However, growing concerns over the incidence of diabetes mellitus have enabled researchers from the other department of Bio-medical science to contribute to the prevention strategies.

This especially stimulated the minds of tissue engineers to work on the novel approach of encapsulation with the objective of providing a better therapy for diabetic patients.

Chang (1964) previously devised a method of developing microcapsules with semipermeable membranes, by setting down polymer around emulsified aqueous droplets using two options as interfacial coacervation and interfacial polycondensation. The researchers of subsequent periods have made this strategy worth fitting for treating disorders like diabetes.

It was reported that islet cells could be better developed by using alginate-poly (L-lysine) alginate membranes (Sun, 1988). They are further described to be biocompatible and viable that would remain impermeable to cells and effector molecules of the immune system.

This could protect the transplanted islet cells against host rejection thus making it a good substitute for immunosuppressive therapy (Sun, 1988). This has strengthened an earlier description of the significance of the bioartificial pancreas. It was described that the transplantation of encapsulated islets could overcome the problems of islet availability and rejection in the treatment of insulin-dependent diabetes with organ replacement (Zekron et al., 1996). The compatibility of the bioartificial pancreas was reported to depend on the reaction of the entrapped islet to the encapsulation technique and material, the reaction of the recipient against the incorporated device ( foreign body reaction), and the reaction of the recipient against the encapsulated islet ( immunology of bioartificial pancreas) (Zekron et al., 1996).

The interactions between a variety of donor and recipient related factors may contribute to certain reactions like inflammation and fibrosis. This study has suggested large experiments on animal models to obtain significant data for the progress in the development of a bioartificial pancreas (Zekron et al., 1996). Encapsulation was believed to allow animal islets or insulin-producing cells to be engineered from stem cells (De Vos, Hamel, & Tatarkiewicz, 2002). As such three major approaches to encapsulation have been focused such as intravascular microcapsules, which are anastomosed to the vascular system as AV shunt and extravascular microcapsules and extravascular microcapsules transplanted in the peritoneal cavity.

Although this intervention was proved to be successful, the extravascular approach was found to be reliable over others since it is associated with fewer complications such as thrombosis and infection (De Vos, Hamel, & Tatarkiewicz, 2002). It was further described that the transplantation of microencapsulated islets close to blood vessels in revascularized solid supports would solve problems concerned with the performance and survival (De Vos, Hamel, & Tatarkiewicz, 2002). Hence, there is a need to implement the strategies found with significant progress for diabetic patients. It was reported that the bioartificial pancreas developed on the basis of encapsulation of islet cells could offer protection to cells from the host’s immune attack. This was mainly for its ability to eliminate the need for immunosuppressive drugs, improved diffusion capacity, and ease of transplantation. Hence, the utility of the Bioartificial pancreas needs to be evaluated.

The material to be used for tissue engineering plays a vital role. Here biodegradable polymers of both natural and synthetic origin were the preferred candidates as they meet the requirements of sutures, scaffolds for tissue regeneration, tissue adhesives, hemostats, and transient barriers for tissue adhesion, as well as drug delivery systems (Nair & Laurencin, 2006). The use of polymers may be largely dependant on the progress achieved in the field of molecular and cellular biology with regard to the development of biotechnological drugs (Nair & Laurencin, 2006). So, there may be a need to take the assistance of a good number of professionals concerned with the relevant field.

Further, the encapsulation technique for treating diabetes may require an evaluation of factors that influence the survival of islet cells. This could in turn rely on the metabolism more probably the internal homeostatic environment that may undergo changes during pathological conditions.

It was described that although encapsulation facilitates the long-term survival of islet grafts in the absence of immunosuppression, it may be confined to only several months due to the effect of nonprogressive pericapsular overgrowth (De Vos et al., 2003). It was revealed that such overgrowth was due to macrophages that produce nitric oxide which, rather than cytokines, may lead to the deleterious effect on the neighboring encapsulated islets. (De Vos et al., 2003). De Groot et al. (2004) reported that the graft function of encapsulated islets may be hindered by a gradual decrease in the glucose-induced insulin response (GIIR), a hyperproliferation of islet cells, and gradual necrosis. This was described to be associated with the increased number of macrophages on the overgrown capsules A concomitant necrosis on the total islet surface was also observed (De Groot et al., 2004intoThe abnormal levels of glucose or the insulin may also indicate an underlying complication that could originate from the cytokines released as a result of altered macrophage function or necrosis.

Although macrophages have an important phagocytic function, their problematic intervention has become a concern in this context. Hence, studies need to be focused on the growth of macrophages that may play a suspicious role during the survival of encapsulated islets.

Further, encapsulated islets may fail to be accepted by the host’s body because of Hypoxia (De Groot et al., 2003). This was revealed when the mRNA expression levels of Bcl-2, Bax, inducible nitric oxide synthase (iNOS), and monocyte chemoattractant protein 1 (MCP-1) were assessed in association with the amount of nitrite and MCP-1 in the culture medium (De Groot et al., 2003). They have described that the increased MCP-1 mRNA levels may serve as an indication that encapsulated islets in vivo contribute to their graft failure by attracting cytokine-producing macrophages (De Groot et al., 2003).

Preventing Hypoxia may significantly ameliorate the function of islet cells and lessen the attraction of macrophages by encapsulated islets (De Groot et al., 2003).

It was reported that the success of islet transplantation would depend on the production of purified alginates of high transplantation-grade quality (Zimmermann et al., 2005).

This would require a designing of immunoisolating alginate-based microcapsules by maintaining the uninterrupted exchange of nutrients, oxygen, and therapeutic factors that are released by the encapsulated cells while simultaneously avoiding swelling and subsequent rupture of the microcapsules (Zimmermann et al., 2005). This strategy would be largely influenced by the implementation of a validated and well-documented technology for cross-linking alginates with divalent cations. As the progress in alginate based therapy was demonstrated by transplantation of encapsulated rat and human islet grafts in diabetic mice models, there may be a scope for diabetic individuals in the near future (Zimmermann et al., 2005). This has indicated the role of encapsulated islet cells that were considered to be immunoisolated. Earlier workers described that immunoioslated islet allo- and xenotransplants, would better have clinical application because of satisfactory findings with alginate microcapsules in both allo- and xenogeneic (porcine) islet transplantation in the spontaneously diabetic dogs and monkeys(Mullen, Maruyama & Smith, 2000). They have also described a reversal of diabetes in mice with cryopreserved, microencapsulated rat islets. Pancreomatized dogs were able to successfully allow the long-term allo- and xenogeneic islet survival of vascularized bioartificial devices (Mullen, Maruyama & Smith, 2000) The limitations of this approach were that a failure of the devices may contribute to thrombosis and the associated problems. This has indicated a partial success of immunoisolated islet transplants in animal experiments. As such, minimizing adverse reactions produced by immunoisolation devices or capsules intended for treating diabetes may ensure the efficacy of technologies concerned with islet encapsulation (Mullen, Maruyama & Smith, 2000)

Recently, it was reported that a new technique known as cell sheet engineering using temperature-responsive culture dishes would facilitate the use of living cells as an immunoisolating membrane (Lee et al., 2008). Since immunisation involves the encapsulation of a graft in a selectively permeable membrane, encapsulation of cellular grafts may protect the graft from immune attack without the need for immunosuppressive agents (Lee et al., 2008). Using the microencapsulation technique, a chondrocyte sheeting immunodelusive immunoisolated bioartificial pancreas (CSI-BAP) was manufactured by means of cell sheet engineering, and an auricular cartilage, which is histologically elastic cartilage from dogs (beagle), was used as a source of immunoisolating membrane. CSI-BAP was able to secrete insulin into the culture medium indicating the efficacy of this approach (Lee et al., 2008). Another recent description has highlighted the importance of a novel composite alginate/poly (lactic-co-glycolic) acid microparticulate system for protein stabilization and delivery using bovine insulin as a model drug (Schoubben et al., 2008). Composite microparticles filled with insulin were reported to show reproducible encapsulation efficiency with a higher soluble insulin content when compared to conventional microparticles(Schoubben et al., 2008). Therefore, it is reasonable to mention that the above strategies may produce significant results if further investigations were made.

Specific Aims

In view of the above information, the main objectives of the proposed study were:

To select a large population of diabetic individuals for testing the efficacy of technology of tissue engineering and encapsulation.
To conduct experiments on animal models.

The rationale for this purpose was that there were poorly available treatment options other than the conventional methods. Pharmacological interventions, gene therapies may accompany the risk of side effects and could not ensure a reliable remedy. In addition, they are not as cost-efficient as the immunization encapsulation technique.

Hence this study was proposed with the anticipation that it could produce better results than might promote a faster drive towards the standard therapeutic establishment of encapsulation.

Research Design and Methods

Specific Aim # 1: To select a large population of diabetic individuals for testing the efficacy of technology of tissue engineering and encapsulation.

Rationale: Reliable treatment of diabetes requires the intervention of novel strategies.

Methods: Online databases would be searched to gather the relevant information. Health authorities would be approached to obtain a permission grant. A survey would be conducted in government and private hospitals to find diabetic patients. Based on the records, a total of 300 patients would be approached and informed about the study. Informed consent would be obtained from patients willing to participate in the study. Those meeting the standard criteria for diabetes classification will be only selected while others are excluded from the study.

Initially, baseline characteristics like age, BMI would be considered. Based on the values obtained, diabetic individuals will be separated into two groups as non-obese and obese.

Fasting blood glucose and CRP levels will be determined. Here, CRP would be used as a concomitant parameter due to the risk of cardiovascular inflammation in diabetic individuals.

Further, islet cell autoantibodies would be determined to detect the defects of islet cells.

Patients who exhibit a markedly decreased pancreatic organ function would be selected for the transplantation experiments. Encapsulated islet cells would be developed by either following the approaches mentioned previously like alginate-poly (L- lysine) alginate membranes, immunoisolated alginate-based microcapsules, or a chondrocyte sheeting immunodelusive immunoisolated bioartificial pancreas (CS-BAP). Encapsulated islet microcapsules would also be obtained commercially to compare the efficacy of the methods developed in the laboratory.

The risk of hypoxia would be monitored by determining the mRNA expression levels of mRNA, Bcl-2, Bax, (iNOS), MCP-1, and nitrite in the culture medium. The risk of macrophage-induced nitric oxide levels and its overgrowth on the neighboring encapsulated islets would be checked by monitoring the cytokine secretions of the macrophage. Human models receiving encapsulated islet cells would be monitored for insulin secretion. These findings would be compared with the strategies implemented for rat models. This is to find the difference in the levels of insulin which might reflect the degree of immune suppression. Obese individuals would also be selected from the 300 diabetic patients for the encapsulation intervention program after conducting preliminary tests on their insulin and glucose levels. Those found with absolutely pancreatic defects would be selected for the surgical removal of the pancreas. Transplantation of encapsulated islet grafts or bioartificial pancreas would be allowed. They would be then monitored for insulin levels. As obese individuals are susceptible to cardiovascular problems, CRP levels would be checked after encapsulated islet cell transplantation. This is to determine whether inflammation would interfere with the survival of islet grafts.

All the subjects will be followed for a period of one year. During this period they could be also evaluated for the negative effects of the host’s immune attack.

Specific Aim # 2: To conduct experiments on animal models.

Rationale: Studies on encapsulation techniques involving animal models are largely needed for confirmation reports.

Methods: An equal number of mice would be selected after obtaining permission from the veterinary authorities. These animal models were initially monitored for their insulin and glucose levels. Those found with a defect in the glucose metabolism would be selected. Those with normal functions would serve as controls.

Pancreas would be surgically removed from the mice and they would be monitored for immune suppression without external treatment. Microencapsulated islet cells obtained previously would be surgically delivered. The mice would be determined for GIIR, the rate of islet cell replication, and islet cell death as described previously (Vos et al., 2004).

The novel approach of CSI-BAP would also be applied. Here, islet cells would be collected and prepared from the rat. CSI-BAP would be cultured for nearly 83 days and the cultured medium would be collected every 24 h to measure the insulin concentrations. The CSI-BAP was examined histologically using hematoxylin and eosin (H&E), and azan dye staining.

Immunohistochemical staining would be additionally performed to demonstrate the insulin production of CSI-BAP. Insulin secretion of CSI-BAP would be observed daily and recorded.

The results obtained on the earlier days would be compared with the later days.

For example, the percentage of insulin secretion obtained on day 10 would be compared with that of day 15. These findings would be later compared with the rats described to be surviving with the defective pancreas. Obese rats would be specially obtained from animal houses of various biomedical research institutions/ laboratories. Their pancreas would be surgically removed and encapsulated islet cells would be transplanted. Immunosuppressive drugs would not be given to human and rat models. The results obtained from the overall experiments would be checked for the approach that has comparatively more significant data.

The treatment option that produced the best results would be further refined by conducting additional studies on both human and animal models. The estimated time period of the proposed project would be fourteen to eighteen months. For the human subjects, six months for the survey and selection, two months for the development and transplantation of encapsulated islet cells, and one-year follow-up. For the animal models, two months for animal selection, two months for the development and transplantation of encapsulated islet cells, and one year follow up.

References

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