My Sci

Well, it’s been quite a while since my last post…again.  Between class and lab I’m pretty swamped these days.  In addition to that, I do put a good bit of work into these posts and I tend to think quality is more important than quantity.  The outcome of which is that I don’t post when I don’t have time to write a good post. 

That all aside, I thought I’d explain the big-picture rationale for one of my projects.  Translation from basic research to clinical application (often referred to as ‘bench to bedside’ is a major goal of the field of biomedical research right now, and for good reason.  The potential implications of my project for patient care are, like many others, several steps removed from the conclusions I hope to draw.  Nonetheless, I think it’s an important aspect of the immune system to understand and one that could lead to changes for the better in therapeutic and preventative medicine.

An immune cell’s ability to move into, out of, and through tissues of the body is essential to its proper functioning.  I’m specifically interested in how T cells accomplish this, but the same principles can be applied to any other immune cell subset.  CD4+ T cells, for example, carry out a number of roles in a variety of tissues depending on the type of immune reaction underway.  In the case of intracellular pathogens like Leishmania major, Toxoplasma gondii or Francisella tularensis, CD4+ T cells need to make direct contact with infected cells wherever they may be.  These pathogens multiply inside host immune cells (often macrophages) where they can evade the mechanisms that phagocytes usually use to dispose of such invaders.  These phagocytes can only do so much on their own.  Without T cells stimulation, specially evolved pathogens can circumvent the limited mechanisms available to a phagocyte on its own.  Once a T cells arrives on the scene, it can stimulate the phagocyte to produce more powerful chemicals and enzymes that help to destroy the pathogen.  A failure in the ability of T cells to respond appropriately to this sort of infection can lead to serious or even lethal complications, as in the case of tuberculosis or visceral leishmaniasis. 

Go home, T cell.  You are drunk.

One critical step in this process is the migration of the T cells from its steady-state location to the site of infection somewhere in the periphery.  This process usually involves many phases that must be executed properly to ensure the efficient clearance of infection.  A naïve T cell under normal conditions can be found circulating through the peripheral lymphoid organs (like the spleen and lymph nodes) searching for antigen presenting cells that are displaying its particular antigen.  Once it finds and APC with its antigen in the presence of inflammatory mediators, the T cells integrates various signals from the environment and matures into an effector cell that is ready to carry out its function.  Next, the T cells needs to leave the lymph node, a process that requires certain surface molecules that it acquired during maturation, and migrate through the lymphatic ducts and into the circulatory system.  Once in circulation, special surface molecules (again, imparted by the maturation process in the lymph node) direct the T cell to the tissue where it is needed.  The T cells must then push its way out of the blood vessel and through whatever interstitium lies between it and the site of infection.  This could mean traversing the alveoli of the lung, the dermis of the skin, or lamina propria of the intestine.  In any case, the T cell needs a specific set of surface molecules that will allow it to migrate efficiently in each of these unique situations. 

Things have been really busy lately; between starting up new projects in the lab and a quick vacation to California.  That, coupled with a lack of inspiration, has resulted in a significant lag in my posting here.  So, in order to get things moving once again, I decided to tackle a topic I’m a little weak on myself: B cells!  And, of course, one can’t discuss B cells without first having a solid understanding of antibodies.

Antibodies are Y-shaped molecules that are either secreted by B cells or expressed on the B cell surface in a form called a B cell receptor (BCR).  Antibodies arise from a process known as ‘VDJ recombination’ in which the heavy chain and light chain which compose the antibody are modified at the genetic level.  When everything goes right, the finished product is a BCR with an entirely unique specificity to some foreign molecule and no reactivity to any self-generated molecules.  When a B cell is activated in a process often, but not necessarily, involving T cells, the B cell integrates signals from multiple sources to determine which antibody class to manufacture.  When a B cell switches the class of its antibody (a process known as ‘class switching’) the antigen specificity is unaltered.  The only change occurs in the end of the antibody that interacts with other elements of the immune system.

Antibodies play many important roles in the immune system and their function is dependent on their particular class.  The different classes of antibodies, or ‘immunoglobulins’ (IgA, IgD, IgE, IgG, and IgM), each serve a unique function in the process of defense against pathogens.  IgE, for example, is most well known for being the culprit behind some common types of allergic responses. This, however, is not IgE’s true evolutionary purpose.  IgE has evolved as a mechanism of defense against large, extracellular parasites (Platyhelminthe worms and nematodes like Trichinella).  IgE functions by being secreted into mucosal tissues (such as in the digestive and respiratory tracts) and attaching to special receptors on the surface of immune cells like mast cells.  These receptors bind to the Fc (fragment of crystallization) portion of the antibody, opposite the Fab (fragment of antigen binding) so that the antigen-specific surface is pointed out and away from the cell.  When the IgE binds its specific ligand, the mast cell is signaled to release granules containing, among other things, histamine.  Histamine is the chemical known to be involved in the vasodilation that results in itching, sneezing, and watery eyes associated with some allergies. However, when this reaction occurs in defense against a parasitic worm, it can result in the pathogen’s destruction or ejection from the body. 

Other antibody types, such as IgG2a and IgG3, serve to activate a class of molecule call ‘compliment’ that can result in the destruction of pathogenic bacteria.  Still others can cover viruses, preventing their entry into cells.  Some can also cover bacteria or cancer cells, signaling macrophages to ingest them.

Antibodies are an elegant and amazing adaptation that is essential to the proper functioning of the immune system (as evidenced by individuals lacking them: http://en.wikipedia.org/wiki/X-linked_agammaglobulinemia)

So, I just officially joined a lab and am starting to plan out potential projects.  One area that my PI is interested in is the CD4+ T cell response to infection by the intracellular protozoan parasite, Leishmania major.  As a consequence, I’ve been doing some research on the subject and thought I’d share and hopefully organize my thoughts a bit better.  Leishmaniasis, the disease caused by parasites in the genus Leishmania is an opportunistic disease, often associated with states of reduced immune function.  As a consequence, leishmaniasis is a major public health concern in poorer countries where people may be living in close proximity, with inadequate sanitation and nutrition.  Leishmaniasis is also often associated with HIV infection and its accompanying immune-deficiency syndrome, AIDS.  L. major itself is a single-celled, eukaryotic flagellate.  It enters the bodies of mammals usually via a vector (sandflies of the genus Phlebotomus in the ‘old world’ and Lutzomyia in the ‘new world’).  Once inside the mammalian host, it is phagocytized into intracellular compartments called phagosomes by macrophages and other phagocytes, though macrophages appear to be L. major’s favorite cell type for invasion.  Normally, this is not an optimal place for a pathogen to be.  Macrophages are particularly good at disposing of pathogens that wind up in their phagosomes.  They are able to produce various enzymes and reactive oxygen species that are toxic to the pathogen which they direct to the phagosomes in order to kill and digest the intruder, though they need T cell help (stimulation) in order to manufacture some of these molecules.  Leishmania, however, employs several mechanisms to avoid this fate which I’ll discuss in a moment, but first: to the immune side of things. 

Under ideal model conditions, the response to an intracellular pathogen goes as follows; antigen presenting cells (APCs) take up antigens (usually proteins made by the pathogen) which they will show to T and B cells to elicit an immune response.  At the same time, APCs like dendritic cells and macrophages are able to sense other pathogen-derived molecules (DNA/RNA, polysaccharides, etc.) using patter-recognition receptors (PRRs; like TLRs and NLRs) to determine what type of pathogen they are dealing with and how the immune system should respond to it.  The APCs then take this information and their antigenic peptides to the lymph nodes where they present it to naïve T and B cells.  When a T or B cell recognizes the antigen, the APC then shares the information it has about the type of pathogen by secreting specific patterns of cytokines which can prime the lymphocytes to exert the appropriate effector function.  In the case of intracellular pathogens, the Th1 type of response (characterized primarily by the cytokine IFN-γ) is the most effective type of response.  Th1 cells (CD4+ T cells that secret IFN-γ) then migrate to the site of infection and stimulate macrophages to produce more toxic molecules and destroy the pathogens that they have phagocytized. 

Usually, this response is effective at preventing serious damage from the invading intracellular parasite.  In some cases, however, a concurrent infection or lack of nutrition can give Leishmania a foothold.  This is where those immune-avoidance mechanisms come in.  L. major has been shown to cleave certain molecules within the phagosomes making it difficult for the host macrophage to present antigen and, therefore, elicit T cell help.  L. major has also been shown, through unknown means, to disrupt the ability of APCs to communicate the type of pathogen to T cells and may even be able to stimulate the release of cytokines that instruct lymphocytes to differentiate into an inappropriate effector type.  Specifically, L. major is able to somehow bias the immune system toward a Th2 (mediated by CD4+ T cells that secret IL-4) type response which is unable to stimulate macrophages or effectively clear the pathogen.  In the worst case, without treatment, this condition can progress to a systemic Leishmania infection and eventually death. 

If I end up with a project related to this subject, I’ll be trying to determine exactly what mechanisms L. major uses to subvert the CD4+ T cell immune response and how that information might be translated into better vaccinations or treatments.  A better understanding of this process may also lead to more effective treatments for other diseases as well; but at the least, should be very interesting.    

Sources:

Fowell and Locksley, 1999 (BioEssays)

Alexander and Brombacher, 2012 (Frontiers in Immunology)

No problem; you post some really great stuff!  It’s cool that you’re interested in studying immunology in grad school.  It’s certainly not for everyone, but I have regretted it for a second.  When I first started searching for graduate programs in immunology, I spent a lot of time on this website: (http://www.gradschools.com/) It may be a bit slanted toward the states, but there seems to be a pretty decent European selection too.  I’ve just finished my first year and officially joined a lab, so my project is only starting to take shape, but I’m interested in T cells and how they interact with other cells under different conditions.  I don’t expect it will be too difficult to work on cancer biology and immunology at the same time.  Research into the relationship between cancer and the immune system is very hot right now.  In fact, the last lab I rotated in is working on exactly that.  The PI in that lab emailed me this (http://www.newyorker.com/reporting/2012/04/23/120423fa_fact_groopman?currentPage=all) which you might find interesting. 

Cancer’s ability to evade and manipulate the immune system has proven to be a major obstacle for biomedical researchers struggling to develop treatment strategies.  As our understanding of tumor physiology has increased, the magnitude and variety of ways in which cancers can influence immune responses has become apparent.  One common primary target for inhibition by cancers, and for study by researchers, is the adaptive T cell response.  The microenvironment generated in tumors is able to suppress T cell responses or bias them toward anergy or tolerance at multiple points (Reviewed in Rabinovich et al., 2007).  Some characteristics of tumors have been shown to favor reduced immune cell recruitment to tumor sites and generation of tolerance.  For example, overactive Stat3 signaling observed in many malignancies leads to the production of vascular endothelial growth factor (VEGF) and IL-10 by tumor cells.  VEGF and IL-10 both produce tolerogenic phenotypes in dendritic cells (DCs) by impairing maturation, NF-κB translocation and, subsequently, upregulation of antigen presentation and co-stimulatory molecules, such as CD80/86 (Niu et al., 2002, Rabinovich et al., 2007, Steinbrink et al., 1997).  This inhibition of DC maturation leads to decreased migration to lymph nodes, reduced capacity to activate T cells, and a tendency to induce anergy to tumor-derived antigens via presentation in the absence of co-stimulation.  As is often noted when describing immunotherapeutic treatment of cancer, some patients benefit more than others.  One suspected culprit of this discrepancy is differential trafficking of effector cells (especially adoptively transferred T cells) to the tumor site related to the expression of chemokines.  The underlying cause of this phenomenon is not well characterized, but it has been observed in many systems (reviewed by Abastado, 2012).  The immunosuppression associated with tumors has also been correlated to the function of regulatory CD4+ T cells (Tregs).  While the precise mechanism of their induction in this system is still unclear, high numbers of Tregs in malignancies is known to predict poor patient prognoses and ablation of Tregs using anti-CD25 monoclonal antibodies significantly increased T cell-mediated tumor clearance (Curiel et al., 2004, Shimizu et al., 1999).   Several tumor tissues have been shown to express B7-H1, a member of the B7 co-stimulatory molecule family.  B7-H1 binds programmed death receptor 1 (PD-1) on T cells and promotes apoptosis, resulting in increased resistance of B7-H1 expressing tumors to T cell-mediated clearance (Dong et al., 2002). 

I've been participating in an ongoing debate with a microbiologist friend of mine.  She loves her pathogens ("bugs," they call them) and thinks their methods of pathogenesis and evasion of the immune system are super awesome.  While I won't argue that host-pathogen interaction is a fascinating topic and a very urgent human health-related concern, I maintain that nothing can mess with the human body quite like the human body. 

Autoimmune and autoinflammatory diseases are some of my favorite topics. (in fact, I'm second author on a related paper that just got published! XD *see source) The ways in which things can go wrong with the immune system are complex and fascinating and the consequences can be devestating. 

IPEX or "immunodysregulatoion polyendocrinopathy enteropathy X-linked sydrome" results from a genetic defect in the Foxp3 transcription factor and is associated with a defect in the function of Foxp3-dependant regulatory mechanisms including the development of regulatory T cells (Tregs).  Mice selectively depleted for Tregs show a massive increase in lymphocyte organ infiltration and pathogenic lymphoproliferation within a matter of a few days resulting in tissue damage and rapid organ failure that closely mimics the pathology of the human disease.  Basically, not a fun time.

Multiple Sclerosis (MS) is an autoimmune reaction against self-antigens from the central nervous system (CNS) resulting in the eventual destruction of neuronal myelin sheaths and other inflammation-associated damage that leads to impaired nervous system function.  Because these antigens are not normally expressed outside the CNS and are, therefore, dependent on Aire-mediated expression in the thymus for tolerance generation, there is a particular degree of leakiness of CNS-antigen specific T cells.  Under homeostatic conditions, this is not a problem; there are ALWAYS autoreactive T cells in circulation, but they are kept in check by many regulatory mechanisms.  However, when these antigens are exposed in an aberrant fashion, resulting from damage due to, for example, viral infection, autoreactive T cells gain access to them and are stimulated by the inflammatory environment to mount a response.  -Of course, this is just one facet of one possible model for MS progression, but it does demonstrate how tentative the balance can be between homeostasis and disease, especially in genetically predisposed individuals.

So, I'm not saying that "bugs" can't do some serious and unpleasant damage (and in terms of sheer numbers, they've got autoimmunity beat) but the point remains that the immune system can keep up with the worst of them.  And there's something so inherently nefarious about attacking from the inside.  I mean, I want to be able to trust my T cells!

I had a interesting lecture today, so I thought I'd type out a bit of what I got from it (doesn't hurt that the final exam is coming up, either 8P).

So, as briefly described in a previous post, central tolerance (ie., the autoimmune-minimizing mechanism of the thymus) is maintained, in part, by expression of self antigen:MHC complexes on the surface of cells in the medulla of the thymus: medullary thymic epithelial cells (mTECs).  Not only do these cells present antigen derived from proteins that one would expect to be present in epithelial cells, but thanks to a molecule called Aire (AutoImmune REgulator) mTECs are also able to express and present many tissue-specific antigens.  These tissue-specific antigens would normally only be encountered in discrete locations in the body (such as the CNS, the eye, or the pancreas) and, therefore, could otherwise present a problem for the prevention of autoimmune reactions.  Since Aire allows for the presentation of these rare proteins in the thymus, the body is able to delete T cells with receptors specific for these self antigens as well.