Natural variation of the bovine lymph node microenvironment and its possible effect on BCG immunogenicity
Led by Dr Lucia Biffar (University of Oxford), with Dr Bernardo Villarreal-Ramos (APHA) and Prof Tracy Hussell (University of Manchester)
Bovine tuberculosis is a disease affecting cattle in the UK and worldwide. It is caused by Mycobacterium bovis, a bacterium closely related to the human pathogen Mycobacterium tuberculosis. The vaccine routinely used to protect humans against tuberculosis is BCG, a live attenuated vaccine developed from a Mycobacterium bovis strain. BCG may also be used as a vaccine against bovine tuberculosis in cattle, however, it is currently only administered in experimental studies. The ability of BCG to induce protection varies considerably between different population groups and amongst individuals from 0-80%, both in humans and in cattle. The reasons for this variability still needs to be understood. After administration a vaccine drains to the lymph node closest to the site of injection. In the lymph node immune cells are trained to recognise the target pathogen, thus enabling them to combat the pathogen during future infection. The lymph node microenvironment is made up of different immune cells and the immune molecules (e.g. cytokines) they express. We suggest that a potential factor influencing the variable efficacy of BCG might be natural variation of the lymph node microenvironment between different individuals. The aim of this project is to analyse the lymph node microenvironment in different cows before and after parenteral BCG administration, by measuring the expression of cytokines and analysing the composition of immune cells. Results from this experiment will provide a baseline for future experiments investigating the impact differences in the lymph node microenvironment might have on the protection conferred by BCG. Furthermore, this baseline data will enable us to design experiments seeking to manipulate the lymph node microenvironment with the aim to improve efficacy of BCG. The results may have wider applicability to other diseases for which vaccines have been difficult to develop.
We have evaluated the immune response of cattle that have been vaccinated subcutaneously or not with BCG in blood and in lymph nodes (LN) before and after vaccination. The LN in mammals is thought to be the site in which the immune response to pathogens is generated and the rational of this project was to determine the natural variation that exists in the lymph node of individuals before and after vaccination with BGC and any possible correlation between this variation and the immune responses detected in peripheral blood. As a group, animals inoculated with BCG showed IFNɣ responses following stimulation with mycobacterial antigens overnight, however, there was animal to animal variation. To evaluate responses in draining lymph nodes, we performed LN aspirates. The amount of cells recovered from LNs varied from animal to animal and additionally temporal variation in the recovery of cells from aspirates within the same animal was observed. We noticed at the end of the experiment, when LNs were harvested, that recovery of LN aspirates by needle puncture (we used a 14” gauge needle which was the smallest bore needle we could use) was inducing relatively large damage to the LN, enough to induce noticeable haemorrhage in the harvested LNs.
Using whole blood and LN aspirate we evaluated the transcription of effector molecules thought to play a role in the immune response to mycobacteria such as IFNɣ, TNFα and perforin, amongst others. We found that there was a trend for a transient increase in the transcription of TNFα in both, blood and LN aspirates. No other difference in any other effector molecule transcript was detected. PBMC samples from these animals have been bio-banked for future experiments.