Characterising the cellular immunity and metabolic response to Mycobacterium tuberculosis and Burkholderia pseudomallei in Indian patients for vaccine design
Led by Prof Chiranjay Mukhopadhyay (Manipal Academy of Higher Education, India), with Prof Susanna Dunachie (University of Oxford, UK) and Prof Mitali Chatterjee (IPGMER, India)
No vaccine is available for the neglected tropical disease melioidosis, and there is need of an improved vaccine for tuberculosis (TB). Both diseases are similar in that the bacteria live inside cells and their defence mechanisms are comparable. People suffering from diabetes mellitus (DM) have a three-fold higher risk of falling sick because of TB, and a twelve-fold higher risk of falling sick because of melioidosis than non-diabetic people. We therefore need to understand the reason why diabetic people are more susceptible to these bacteria. Likewise, understanding the response from the immune system of diabetic patients suffering from TB or melioidosis, will help in developing a new or improved vaccine.
We will combine expertise at the Department of Microbioloy at the Manipal Academy of Higher Education (MAHE) in India and the University of Oxford, to establish how the metabolic status of immune cells differ in diabetic people that makes them susceptible to TB and melioidosis. In particular, we will investigate the metabolism of immune cells of these patient groups. Just like every other cell in the human body, immune cells need energy to function, especially when they have to be very active, as during an infection when they have to 'fight' the pathogen. Immune cells mostly use sugar (glucose) to acquire their energy, but the cells can use different pathways to convert glucose into energy. We will investigate how the metabolism of immune cells of diabetic patients responds to an infection with TB and melioidosis and if this differs from non-diabetics. We believe this will help to design better vaccines for TB and melioidosis in people with diabetes, allow development of better immune correlates of protection, and can also help us develop ways to use medicines at the time of vaccination to boost immune responses in people with diabetes.
The main goal of our project is to establish how the immunometabolic status of the diabetic people make them more susceptible to tuberculosis.
(TB) and melioidosis diseases in Indian subjects. In this regard, the first study objective includes understanding the cellular immune response to specific target antigens with respect to TB and melioidosis diseases that will help in getting a better insight into determining which antigens are immunogenic and which immune responses are upregulated and protective. This is followed by targeting the study towards metabolism of different T-cell population, the main players of the adaptive immune system, which will aid in obtaining insight into understanding the difference in the metabolism of T-cells in each patient groups, and examination of the interaction between T cell response and metabolism will illuminate potential mechanisms for immune impairment in diabetes leading to susceptibility to these infections. Finally, we will correlate the immunometabolic dysfunction to increased susceptibility towards TB and melioidosis.
We have standardized peripheral blood mononuclear cells (PBMC) isolation and whole blood stimulation assay protocols. The blood samples of all the 9 recruited patients in our study have been processed for PBMC isolation and whole blood stimulation followed by cryopreservation until further downstream processing is performed. Initial standardization of ELISpot assay was performed using cryo-revived PBMCs’ derived from healthy individuals. The standardized ELISpot plates will be couriered to nearby centre for further analysis.
We will be carrying out ELISpot assay on the isolated PBMCs from the patients recruited for the study followed by further analysis of the plates based on the ELISpot readings obtained. Standardization of intracellular cytokine staining experiment will be performed prior to the patient sample processing for the same. Study of T-cell metabolism during infection and recovery will be completed using flow cytometry and Seahorse Extracellular Flux analysis. Our work will lay the foundation for the identification of drug candidates for protection in diabetes mellitus (DM) and drug discovery for possibilities of clinical interventions towards cure of TB and melioidosis.
Standardization of immunometabolism experiment protocols are on the way preceding its application on the blood samples of the recruited patients.
Antigen specific cell responses will be determined using flow cytometry. Study of T-cell metabolism during infection and recovery will be completed using flow cytometry and Seahorse Extracellular Flux analysis. Our work will lay the foundation for the identification of drug candidates for protection in diabetes mellitus (DM) and drug discovery for possibilities of clinical interventions towards cure of TB and melioidosis. Our studies will help understand the dependency/preference of nutrient availability and subsequent immunometabolic changes for the Bp infection progression. It will also help in understanding the increased susceptibility to infection that is a notable concern for diabetic patients. Studies from patient samples and cell line work will lay the foundation for more mechanistic studies that remain unexplored.