A role for the human mesenchymal stem cell secretome in attenuation of cytokine-induced apoptosis in pancreatic beta cells

Abstract

Diabetes is a lifelong condition caused by an inability of the body to break down glucose due to a defect in either insulin synthesis or the target cells becoming resistant to secreted insulin. There are two main types of diabetes, type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus. T2DM mainly occurs due to insulin resistant, inability of cells to respond to normal levels of insulin, the treatment of T2DM usually involves exercise, diet control, drugs and in some cases insulin is needed. In T1DM, the immune system starts to attack the ß-cells, the cells responsible for insulin hormone synthesis and secretion from the endocrine pancreas. The aim of T1DM management is to restore carbohydrate metabolism as close to the normal condition as possible. To achieve this goal insulin hormone must be provided daily. Insulin is given in different ways such as injection (which is the most common method), pump and inhalation. Insulin sources can be either recombinant or animal-based. Treatment of T1DM is continuous and even with the best treatment options people with diabetes can develop serious complications such as acute diabetic coma or other long-term complications. These include diabetic cardiovascular disease, diabetic retinopathy, and diabetic nephropathy. Injection of insulin and daily monitoring of glucose levels presents a substantial burden to both the diabetic patient and also to associated dependents or carers i.e paediatric diabetes. So there is a strong need to develop an alternative treatment to reduce the burden, enhance control, and even ultimately cure diabetes mellitus.

A recent medical invention is the use of stem cells in the treatment of many disease conditions. Stem cells have a remarkable ability to develop into many different cell types. Stem cells therapy offers a new method for treating disease such as diabetes. The results of many studies demonstrate the capability of mesenchymal stem cells (MSCs) in the treatment of bone disease, cardiac disorder, and multiple sclerosis. However, much work remains to be done in the clinic and laboratory to optimise the use of these cells.

The main aim of this study was to explore the therapeutic effectiveness of MSCs conditioned media to restore the viability and function of ß-cells.

In order to establish an in vitro model of cytokine driven ß-cells apoptosis, pancreatic ß-cells were treated with rising concentrations of pro-inflammatory cytokines TNF-a, IFN-? and IL-1ß and endotoxin LPS for 24 h. The optimal concentration of each cytokine or endotoxin was assessed by MTT assay. Optimal concentrations were deemed to be those that induced an approximate reduction in cell viability of 50%. The cells were treated with the optimal concentration and cell viability was monitored over time in addition to assessment of anti-apoptotic gene induction via qPCR assay. Mesenchymal stem cells (MSCs) secretome was collected as conditioned media and the ß-cells cultured in non-conditioned and conditioned media during cytokine-driven apoptotic induction. ß-cell viability and anti-apoptotic gene expression was determined to evaluate the therapeutic effectiveness of mesenchymal stem cells conditioned media (MSC-CM) in protecting ß-cells from pro-inflammatory cytokines.

We observed a significant increase in the viability of pancreatic ß-cell lines cultured in conditioned media when compared to those cultured in non-conditioned media. After that, we sought to identify a possible candidate that is present in the MSC-CM and help the cells to overcome the effect of pro-inflammatory cytokines. We found a high concentration of IL-10 in our conditioned media, in addition to the presence of IL-4, PIGF and VEGF in variable amount. Based on our present findings, cytokine-induced apoptosis is mediated through the TRAIL-dependent pathway. However, the addition of MSC-CM blocked cytokine-induced apoptosis and downregulated the genetic expression of A20 and TRAIL. Also, IL-10 was able to block IFN-? and TNF-a-induced apoptosis.