Molecular Basis of the Disease state

COPD is a complex condition that has only recently been studied on the molecular level. Although ~95% of COPD is thought to be caused by cigarette smoke, the exact cause-and-effect nature of the pathophysiology of COPD is not well-understood. Nonetheless, several correlations have been drawn, and several “pieces of the puzzle” have been studied fruitfully.

Genetic Causes: Alpha-1 Antitrypsin Deficiency

Alpha-1 antitrypsin (A1AT or AAT) is a serpin and plays an important role in keeping serine proteases such as neutrophil elastase in check during inflammation in order to minimize damage to lung tissue. Mutations in the SERPINA1 gene causes A1AT deficiency or a nonfunctional form of A1AT. The Z variant of this gene is the most common, and yields the Z variant of the A1AT protein, with a Glu342Lys mutation (Jezierski 2001).

The wild-type M variant A1AT protein has a reactive loop that is responsible for proteinase inhibition. The mobility of this loop is controlled by electrostatic interactions between the side chains of Glu342 and Lys290, which are located at the junction of the reactive loop and a beta sheet. The Glu342Lys mutation (“the Z mutation”) introduces instabilities in the reactive loop region, decreasing its efficacy as a protease inhibitor (Jezierski 2001). The effect of this mutation on the structure of A1AT is shown in Figure 5.

BCM441 - a1at mutation structure
Figure 5 – A) Wild-type A1AT with favorable side-chain interaction between Glu342 and Lys290. B) ‘Z’ mutant A1AT with unfavorable replusion between positively-charged Lys-NH3+ R groups. (Original figure using Swiss PDB Viewer & PDBID: 1KCT)

 

Reactive Oxygen Species

Reactive Oxygen Species (ROS) are produced by inflammatory cells. Although the production of ROS is a potent defense mechanism, it can also cause damage to lipids, proteins, and DNA, which is thought to be correlated to lung tissue damage in COPD cases.

ROS can react with arachidonic acid to form 4-hydroxynonenal (4-HNE), an α,β-unsaturated hydroxyalkenal, which plays a key role in signal transduction in a variety of pathways. In a 2012 study by Takimoto et al, it was found that exposure of mice lungs to 4-HNE induced the accumulation of inflammatory cells, “enlarged the airspace, and induced goblet cell metaplasia [replacement of differentiated cells] of the airways”, which are characteristic COPD physiology (Takimoto 2012). Structures are shown in Figure 6.

BCM441 - HNE
Figure 6 – Structures of Arachidonic acid, 4-hydroxynonenal (Original Figure)

 

NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) is a protein complex that controls transcription of DNA, which is involved in cellular responses to various stress stimuli. Although it has many functions and is involved in many processes, cytokine production is its primary relevance to COPD. ROS activate NF-κB, which further amplifies the inflammatory response. The exact pathway of this activation is unknown, but there are several redox-sensitive steps in the activation pathway that are known (Barnes 2003).  Because inflammatory cells can release ROS, there is potential for positive-feedback in this process, which can yield significant damage to lung parenchyma due to ROS and elastase activity.

Matrix Metalloproteinases

There is increased macrophage expression of matrix metalloproteinase 1 (MMP-1, collagenase) and MMP-9 (gelatinase B) in COPD patients. In a 1997 experiment, it was demonstrated that emphysema induced by chronic cigarette exposure is prevented in MMP-12-/- mice (Hautamaki 1997) and in two 2000 studies, it was shown that in MMP12-/- mice, emphysema induced by interleukin 13 (IL-13) and interferon- γ (IFN- γ) overexpression is reduced, as well as a reduction of recruitment of monocytes into the lung. It is speculated that MMPs activate the latent form of TGF‐β (transforming growth factor beta, a cytokine), resulting in the recruitment of monocytes (Barnes 2003). This possible mechanism of action is shown in Figure 7.

BCM441 - MMP12 figure
Figure 7 – Schematic of MMP-9 activity on TGF-β, A1AT, neutrophils, and downstream generation of COPD symptoms. (Source: Barnes 2003)

 

The structure of MMP-12 has been resolved (Nar 2001), and its mechanistic activity has been elucidated. The protein binds a zinc ion and three calcium ions, and has an N-terminus proenzyme domain that is cleaved, activating TGF‐β.

Epigenetic Factors

Cigarette smoke has been shown to impair the activity of HDAC (histone deacetylase) in macrophages, which has been correlated to amplification of the expression of inflammatory genes (Barnes 2003).

Summary of Molecular Basis

The inflammatory response, when uncontrolled, leads to the pathophysiology of COPD. Neutrophils release elastase as part of the inflammatory response, which can damage lung tissue to great extents if the individual is A1AT-deficient. ROS can affect several types of biomolecules; one such target is arachidonic acid, which can be converted to 4-HNE, which can induce the recruitment of inflammatory cells. Matrix metalloproteases, similarly, activate TGF- β to recruit monocytes, which can damage lung parenchyma. Lastly, cigarette smoking can affect the activity of HDAC, and amplify the expression of inflammatory genes in an epigenetic fashion.

6 Replies to “Molecular Basis of the Disease state”

  1. Hi Besher. Thanks for your information. I have a few questions – 1) When you discuss the E342K mutation antitrypsin deficiency as being a less efficacious protease, do you mean that this mutation results in a lower binding affinity for the protein substrate? 2) Regarding the epigenetics portion, you mentioned that smoking impairs the activity of histone deacetylase and amplifies the expression of inflammatory genes. Usually, acetylation loosens the chromatin structure and results in higher levels of expression, and deacetylation tightens the chromatin structure and results in decreased expression. Do you have any thoughts about why this seems to be contradictory? 3) Are there any other effects of ROS interacting with arachidonic acid? We learned in BCM441 that eicosanoids like arachidonic acid are involved in many cellular processes, so perhaps this has more global implications than just COPD. I really enjoyed your page – thanks!

    1. Hey Zach,
      1) You’re right in saying that the E342K mutation causes a decrease in substrate binding, I’ll re-word that, thanks!

      2) You are of course right in terms of the effects of acetylation and deacetylation of histones and their effects on gene expression. If deacetylation decreases gene expression, then the impairment of deacetylation will increase gene expression. Here’s part of a discussion from the original paper in 2001: “We have shown that cigarette smoking is associated with a reduction in HDAC2 expression and
      activity in bronchial biopsies and alveolar macrophages. This finding is correlated with an
      increase in basal and IL-1β–stimulated TNF-α expression in alveolar macrophages”. I think the wording might have confused you 🙂

      3) A 1999 JBC paper studied the effects of NO2 reaction with arachidonic acid, and they found the products to be “a mixture of arachidonic acid isomers having one trans-bond and three cis-double bonds”, which they termed “iso-eicosanoids”. From what I could find, they are termed “biologically-active”, but their effects on the body are not understood. They are, however, used as a measure for oxidative stress in the body, which makes sense. Good questions!

  2. Hi Besher, thanks for your report on COPD. I was intrigued by your section on oxidative stress and its effects on COPD presentation because oxidative stress was also a part of my project. It was also interesting because of the way the immune system was involved, in that there is an uncontrolled inflammatory response, but never any intervention from the adaptive immune system, unlike autoimmune diseases. I was wondering if any of your research pointed in the direction of mutations in anti-oxidant enzymes. While you did mention the possibility of a positive feedback loop in COPD involving ROS, is there a possibility that individuals who have mutations in anti-oxidant enzymes will be more susceptible to developing COPD either naturally or through cigarette smoking? Lastly, is the A1AT mutation listed in your opening paragraph related to genetic disposition or is it the effect of cigarette smoking? Or is it both? Thanks for the interesting read!

    1. Hey Matt,

      Good questions! There is a 2007 paper that studied the activities of anti-oxidant enzymes in patients with COPD. The abstract beings with: “Among the 42 studied candidate genes, the expressions of mRNA for catalase, glutathion S-transferase P1 (GSTP1), glutathion S-transferase M1 (GSTM1), microsomal epoxide hydrolase (mEPHX) and tissue inhibitor of metalloproteinase 2 (TIMP2) were significantly decreased in COPD lung tissues compared with those in non-COPD tissues, and most of these decreases were significantly correlated with the degree of airflow limitation.” COPD is a complex disease with a lot of factors to consider, and enzyme activity is certainly one of them, as this paper shows (in addition to A1AT activity, and smoking, certainly).
      In terms of the A1AT mutation, as far as I have read, that is simply a genetic factor that has not been shown to be affected by environmental factors. Thanks for reading!
      Source: http://www.sciencedirect.com/science/article/pii/S109455390600071X

  3. Great work on your article! I like how you discuss various factors in COPD, until now I would have thought that it occurs solely through smoking related complications.

    Arachidonic acid has been associated with airway fibrosis and prostaglandin mediated inflammation. However in terms of abundant oxidative stress it can also augment the molecule such that it goes through a more powerful inflammatory signaling via lipooxygenases/leukotrienes. Do you know if those who are more susceptible to COPD preferentially signal through the luekotriene route?

    1. Hey Dana,

      Thanks for your kind words, I’m glad I’ve contributed to your knowledge!

      I did come across a paper that studied the possible shifting in metabolism of arachadonic acid from anti-inflammatory lipoxonis pro-inflammatory leukotriene B4, which was indeed what they found, in patients with advanced COPD. I’ve mentioned this in a few other comments: I personally view COPD as a rather complex and comprehensive disease, that involves many different factors all at once – which is potentially troubling, as sometimes, the biochemistry is not necessarily inter-related, with one cause (which could make treatment very difficult, if not impossible). However, understanding these underlying causes is very important for the future of treatment and management. Thanks again for your comment!

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