2 Microarray Analyses of Inflammation Response of Human Dermal Fibroblasts to Different Strains of B. burgdorferi S.S.

This interesting abstract just got posted on PubMed and is in PLoSONE:

Microarray Analyses of Inflammation Response of Human Dermal Fibroblasts to Different Strains of Borrelia burgdorferi Sensu Stricto

Schramm F, Kern A, Barthel C, Nadaud S, Meyer N, Jaulhac B, Boulanger N.

Abstract

In Lyme borreliosis, the skin is the key site of bacterial inoculation by the infected tick, and of cutaneous manifestations, erythema migrans and acrodermatitis chronica atrophicans. We explored the role of fibroblasts, the resident cells of the dermis, in the development of the disease.

Using microarray experiments, we compared the inflammation of fibroblasts induced by three strains of Borrelia burgdorferi sensu stricto isolated from different environments and stages of Lyme disease: N40 (tick), Pbre (erythema migrans) and 1408 (acrodermatitis chronica atrophicans).

The three strains exhibited a similar profile of inflammation with strong induction of chemokines (CXCL1 and IL-8) and IL-6 cytokine mainly involved in the chemoattraction of immune cells. Molecules such as TNF-alpha and NF-κB factors, metalloproteinases (MMP-1, -3 and -12) and superoxide dismutase (SOD2), also described in inflammatory and cellular events, were up-regulated.

In addition, we showed that tick salivary gland extracts induce a cytotoxic effect on fibroblasts and that OspC, essential in the transmission of Borrelia to the vertebrate host, was not responsible for the secretion of inflammatory molecules by fibroblasts.

Tick saliva components could facilitate the early transmission of the disease to the site of injury creating a feeding pit. Later in the development of the disease, Borrelia would intensively multiply in the skin and further disseminate to distant organs.

Link: http://www.ncbi.nlm.nih.gov/pubmed/22768217

Comments:

Take note of that last paragraph:

"Tick saliva components could facilitate the early transmission of the disease to the site of injury creating a feeding pit. Later in the development of the disease, Borrelia would intensively multiply in the skin and further disseminate to distant organs."

Do you think the implications of the above fit in nicely with the mathematical modeling of Borrelia burgdorferi infection cycles mentioned in an earlier entry?

Why or why not?

See:

Abstract: Population Dynamics Of Borrelia burgdorferi In Lyme Disease
http://campother.blogspot.com/2012/04/abstract-population-dynamics-of.html

The implications - for me at least - seem to fit a model where the first wave of infection dies off but then a bigger, immune-resistant subpopulation explodes onto the scene (the site of infection).

Awaiting PLoSONE to publish the full text so I can give a more thorough analysis...

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

10 Commentary On Chronic Lyme Disease and Skepticism

When I began writing this blog, I originally wrote it to give a point-by-point analysis of how one article in the Chicago Tribune, "Chronic Lyme: A dubious diagnosis", did not address the issue of whether or not Lyme disease could be a chronic and persistent infection after initial or delayed antibiotic treatment. I wrote the analysis using Carl Sagan's Baloney Detection Kit, because I thought it was a good framework for deconstruction of the article.

I thought this analysis would attract the attention of a number of skeptics, and from there, we'd engage in a lively discussion about how the article skirted the issue of whether or not chronic Lyme disease is possible and dive into the evidence for and against chronic Lyme disease being a persistent - if only subclinical - infection.

It never happened. 

Over a year later, and it still didn't happen.

And, to this day, only one person has ever left me a comment which directly challenged what I was writing about and judged the content of this blog - at least as far as the idea of persistent infection is involved.

That comment was this:

You error in giving any credit to the "chronic lyme disease" movement.

I recommend this paper in the current issue of The Lancet (Sep 2011;11:713-719) :

Antiscience and ethical concerns associated with advocacy of Lyme disease

Abstract: Advocacy for Lyme disease has become an increasingly important part of an antiscience movement that denies both the viral cause of AIDS and the benefits of vaccines and that supports unproven (sometimes dangerous) alternative medical treatments. Some activists portray Lyme disease, a geographically limited tick-borne infection, as a disease that is insidious, ubiquitous, difficult to diagnose, and almost incurable; they also propose that the disease causes mainly non-specific symptoms that can be treated only with long-term antibiotics and other unorthodox and unvalidated treatments. Similar to other antiscience groups, these advocates have created a pseudoscientific and alternative selection of practitioners, research, and publications and have coordinated public protests, accused opponents of both corruption and conspiracy, and spurred legislative efforts to subvert evidence-based medicine and peer-reviewed science. The relations and actions of some activists, medical practitioners, and commercial bodies involved in Lyme disease advocacy pose a threat to public health.

And pertinent to this blog, the authors conclude, in part:

"This situation is not likely to end anytime soon. As with other antiscience groups, many Lyme disease activists are well funded and often connected to influential political and media sources. Treatment of Lyme disease with long-term antibiotics is profitable for LLMDs and can be falsely reassuring to patients, who believe that they have a debilitating chronic infection and thus do not seek diagnosis and treatment for other disorders. There is no deficiency of either new patients or activists. The medical anthropologist Sharon Kaufman wrote that “Information technology has transformed the way trust and knowledge are produced”.67 Most people now find medical information on the internet, and the websites of LLMDs and activists are often viewed as legitimate and reliable sources of information, which they may not be."

         - LindaRosaRN


I responded to her, asked her a few questions, and she never responded in return. A drive by. 

She obviously wanted to be heard but did not want to engage. Her comment was on a post about the Huffington Post and Under Our Skin; it wasn't even related to the antiscience letter to which she referred.

And since then, it's been crickets all around when it comes to skeptics  - even though I get visits from Spirochetes Unwound (of Research Blogging), and other web sites which are science and skeptic-oriented on a number of levels.

It's almost as if someone gets to the front door of this site and decides it's not for them just because it mentions the words, "chronic Lyme disease".

I'm guessing the skeptical person who stumbles upon this page with that first glance thinks, "Oh, well, it's another one of those chronic Lyme activists. Why are they ignoring all the evidence that after antibiotic treatment, those with persisting symptoms develop an autoimmune disorder? I'm getting out of here. I'm not wasting my time on this..."

If they're thinking that, then perhaps an overhaul is in order for this blog because I'm not trying to ignore the evidence about autoimmunity or immune dysregulation related to persisting symptoms.

I'm trying to look at a number of different hypotheses - including chronic infection - because these topics are of interest to other patients and I think they all deserve discussion. And there are other hypotheses outside of chronic infection and autoimmunity which are being researched such as chronic Lyme disease as a metabolic disorder, and I give them exposure.

Because I enjoy science, I look at research and science news related to tickborne infections and their reservoir hosts in general, without any particular goal in mind other than to say, "Hey, look at this - isn't this weird/interesting/informative?" I want to inspire the curiosity of the world I have in others.

At first glance, this blog has been primarily written for other patients with persisting symptoms related to Lyme disease infection. But it has also been written to open the floor to anyone for discussion about different causes for persisting symptoms.

This includes skeptics. They are welcome to comment here and share their perspective, as long as they aren't rude and can state their position while providing supporting evidence.

After all - I have a history of being a skeptic myself, and at times don't just want the cheering section behind me saying they support me as a fellow patient. I also want someone to occasionally shake me out of my complacency if I've been stuck in it. I occasionally want someone to present me with a different point of view or new evidence if they have it so I can consider other lines of inquiry in trying to understand my condition.

I have to wonder sometimes if I need a colorful banner or animated message on the top left corner of the homepage which states, "I am a skeptic and a patient with persisting symptoms related to Lyme disease".

Or maybe I need a paragraph at the top of one of my sidebars in order to say more about who I am.

Something like this:

Side note to all scientists and skeptics, in case you missed it: 

This blog is written from the perspective of one person who has been in the unfortunate position of being a skeptic who is suffering from persisting symptoms after receiving an infected tick bite.  

I want answers to questions such as, "What on earth does someone in my position do?" and "What kind of pragmatic approach can I have to dealing with my illness when I am a skeptic and want evidence to support my position and the decisions I make?" and "What research is out there which supports one or another position as to the cause for my condition?"

This has to be done carefully, though, for the sake of my audience.

While I want skeptics to feel welcome here,  I don't want to scare other patients away who are dealing with the same struggles. And I want everyone to feel welcome here and not like they have to constantly defend their illness to others who may not only be skeptical - but who have the potential to engage in less than civil behavior online.

My concern here is not unwarranted. I have read skeptic sites for a number of years, and while I can read sites where issues concerning chronic Lyme disease are debated by infectious disease specialists, researchers, doctors, and patients - such as this BMJ series of Rapid Responses - I practically choke on some of the comments on this chronic Lyme disease post on Respectful Insolence blog, because some of the exchanges are insulting and do little to advance serious discussion about Lyme disease. (Some of them are thoughtful and intelligent, too, though, so I don't avoid reading the thread entirely just because there are insults.)

And I have to remind myself that any blog on the internet has the potential for ill-informed comments, regardless of the original content.

One response on the above linked blog betrays a certain basic ignorance about Lyme disease:

"Yikes, always knew Chronic Lyme disease was complete woo. It's called an anxiety disorder, get your meds. Even acute Lyme disease is looking to be a bit far fetched, now I am far from an expert but it seems a little bit out of a sci fi novel for a tick to harbor some strange virus, who proved this and what experimental data did they have? What were the experiments that prove that some tick borned virus is the cause of a human disease in the first place?"

What the... 

Acute Lyme disease looks far fetched and it must be sci fi that a tick harbors a strange virus?

Far from expert?

Indeed. 


And some comments are thoughtful and the response is respectfully delivered even if the content is something a number of patients would find disagreeable:

"...I don't doubt your story, but given how common tick bites are, it seems likely that an association could be found regardless of whether or not the tick bite actually caused the condition. Thus, anecdotes are not that useful in understanding the disease. Anecdotes don't tell us cause and effect. They show us why we should care, and they are important for that reason, but they do not tell us what is true. It takes a lot of carefully collected data to do that. 

How do you know the antibiotics are really responsible for your improved state of health? Remember, coincidences do happen -- during the summers I take Pulmicort, but is it really making my asthma better, or is the variable course of my asthma just being kind to me right now? I have anecdotes galore, but it's the clinical trial data that convinces me the Pulmicort really is working and it's not just my own desire for it to work making it seem as though it is.

Indeed, one day the truth may come out. But it'll have a hard time coming out when the science of Lyme disease is decided by politicians, and when those most invested in chronic Lyme disease would rather spend their effort treating patients than finding out whether or not they're actually doing the right thing for their patients. Too many of the positive studies I have seen have been enterprises in proving a point rather than testing an hypothesis; science can't be fair or unbiased or *truthful* if it does not entertain equally the possibility of being wrong."

They have a valid point. I can see that.

The problem is, when is this "one day where the truth may come out"?

I have to make decisions about what to do about my health TODAY.

That's the one truth which few of these exchanges seem to address:

How does one make an informed decision as to how to approach and treat one's condition when there is a medical controversy over how to treat it where one of the two options is not to treat it at all - and doing nothing leads to more suffering?

While the author takes the time to explain that anecdotes are not data (with which I agree) and that tick bites in and of themselves are common, there is plenty of medical research and documentation which provides evidence that a subset of infected Lyme disease patients go on to have persisting symptoms after initial treatment. For some of these patients, these symptoms are severe. For a number of them, symptoms persist not for months but for years.

The IDSA and other medical institutions and researchers acknowledge this condition. It's not well understood - but this phenomenon has been documented and is not the creation of a handful of woo charlatans looking to cash in on me.

And I am, for whatever unfortunate reason, the poor bastard who has this condition. And I:
- had a documented tick bite,
- developed an EM rash,
- received a positive blood test later on,
- developed all these symptoms which were clinically diagnosed as Lyme disease.

Unlike the claim made by Ms. Rosa above, no one has tried to con me into this diagnosis. I had a textbook case of Lyme disease. The evidence was there.

If anything, when I first contracted Lyme disease, I thought I was getting it taken care of early enough and would not have to touch the "Lyme controversy" with a twenty foot pole.

Unfortunately, life had other plans for me.

I ended up not being able to work, got sicker than I'd ever been in my life for months on end, then finally got better and off antibiotics and return to work... only to relapse, have to leave my job, and try to figure out what to do next.


Why me? I didn't ask for this. As someone who reads Free Inquiry on occasion, worked as a researcher, lived in a dorm full of physics and chemistry geeks, and thought Carl Sagan was one of the most brilliant and compassionate minds on Earth, I really did ask "Why me?".

But you know, the answer to a pathetic "Why me?" really is "Why not me?". No one asks for this. No one asks to be hit by a car, abused as a child, diagnosed with breast cancer, or to watch a parent die. We are all subject to things which happen beyond our control. It was just the random luck of the draw that someone who happened to be a skeptic would get a condition of which others are skeptical.

Ticks don't care if you like Sagan or vote for Romney. They just want lunch.

For better or worse, here I am. Now, how can I make the best of this? What can I make of this situation now that I'm smack in the middle of it?

I've thought that all I can do is write, research, and encourage others to do research. Learn and encourage others to learn more about tickborne illnesses. Encourage those who are out there who are working in microbiology and biology to engage in education and outreach which helps patients and sidesteps the politics. Educate one doctor at a time that someone with my condition exists, and that a tickborne infection can lead to chronic symptoms. Warn others and tell them to protect themselves when they go hiking and camping so they can avoid my fate.

Mainly what I want to do is get better and get back to living life as I was before - but this time, perhaps a little humbler, a little wiser, and with more compassion for others and more gratitude for myself.

In meantime, I keep hoping I'll meet others here who came from a similar background.

So far, in terms of scientists, I have had a few other Lyme disease patients with science degrees and research positions (or friends and partners of patients with science backgrounds) comment on this blog. Outside of these patients, researchers have commented less often and anonymously.

And I keep hoping someone who is a skeptic and also struggling with chronic Lyme disease or perhaps CFS/ME will come forward and we can review research together and try to make sense of it and figure out what the best course of action is. Even better? I keep hoping to see a microbiologist who researches Borrelia burgdorferi and has had persisting symptoms from Lyme disease take up blogging on their own and publish more insights of which are but a faint glimmer in my own head.

But so far? Crickets.

Surely we can do better than this. Surely one can light more candles against the darkness?

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

3 Abstract: Population Dynamics Of Borrelia burgdorferi In Lyme Disease.

Credit goes to Joanne, of the Looking at Lyme blog, for mentioning this abstract: It describes a response to Borrelia burgdorferi infection in mice where the first immune response almost clears the infection - but approximately 1 week post infection, the bacterial population recovers and reaches an even larger size before entering the chronic phase.

Front Microbiol. 2012;3:104. Epub 2012 Mar 22.
Population Dynamics of Borrelia burgdorferi in Lyme Disease.
Binder SC, Telschow A, Meyer-Hermann M.

Source
Department of Systems Immunology, Helmholtz Centre for Infection Research Braunschweig, Germany.

Abstract

Many chronic inflammatory diseases are known to be caused by persistent bacterial or viral infections. A well-studied example is the tick-borne infection by the gram-negative spirochaetes of the genus Borrelia in humans and other mammals, causing severe symptoms of chronic inflammation and subsequent tissue damage (Lyme Disease), particularly in large joints and the central nervous system, but also in the heart and other tissues of untreated patients.

Although killed efficiently by human phagocytic cells in vitro, Borrelia exhibits a remarkably high infectivity in mice and men. In experimentally infected mice, the first immune response almost clears the infection. However, approximately 1 week post infection, the bacterial population recovers and reaches an even larger size before entering the chronic phase.

We developed a mathematical model describing the bacterial growth and the immune response against Borrelia burgdorferi in the C3H mouse strain that has been established as an experimental model for Lyme disease.

The peculiar dynamics of the infection exclude two possible mechanistic explanations for the regrowth of the almost cleared bacteria.

Neither the hypothesis of bacterial dissemination to different tissues nor a limitation of phagocytic capacity were compatible with experiment.

The mathematical model predicts that Borrelia recovers from the strong initial immune response by the regrowth of an immune-resistant sub-population of the bacteria. The chronic phase appears as an equilibration of bacterial growth and adaptive immunity.

This result has major implications for the development of the chronic phase of Borrelia infections as well as on potential protective clinical interventions.

(Special thanks to Frontiers in Microbiology journal for having what appears to be a solid peer review process and Creative Commons license.)

Comments:


I can't wait to read the full text of this paper. This is an intriguing abstract and it leads to more questions.

How did these researchers come up with their mathematical model? Have other researchers previously observed this second wave of bacteria during infection? What about evidence of peaks in immune response in Borrelia burgdorferi infected animal models which have been documented?

In which way, precisely, is the regrowing immune-resistant sub-population actually immune-resistant? What is happening to B cells and T cells in relationship to this second phase of Borrelia?

From Fig. 3 of Tunev et al., 2011.  Day 8 of infection.  The arrows point to intact extracellular B. burgdorferi in the subcapsular sinus of the lymph node, which was culture positive beginning on day 1 of infection.   Source

Do these phases respond with plasma B cells containing low quality antibodies in germinal centers?

Is there typical somatic hypermutation and antigen affinity or not? What is happening to the T-cell independent response?

And does this model have any implication for antibiotic treatment? As in: Does this second immune-resistant sub-population also have a different response to antibiotics than the initial wave? This model is about infection without treatment, and it is not discussed what the implications could be.

Lots of questions here...


UPDATE: Joanne has informed me the free full text is available online. See: http://www.frontiersin.org/Microbial_Immunology/10.3389/fmicb.2012.00104/full

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

7 Did Isabel Diterich Have The Cure For Chronic Lyme Disease?

One researcher whose papers I've been reading recently is Isabel Diterich's. Several years ago she published two papers on Lyme disease which grabbed my attention because they not only revealed a hypothesis of immunosuppression caused by Borrelia burgdorferi spirochetes - but they also revealed a potential cure for chronic Lyme disease.

I say "potential" here with this caveat:

While the treatment did appear turn a man who was disabled into what sounds like the picture of health for at least eight years, he had to take filgrastim for almost two weeks. And filgrastim is an immune modulating drug which can have serious side effects in some people - there have even been a few fatalities.

However, most of the people who have suffered serious side effects from filgrastim were cancer and leukemia patients who already had serious health problems and were at greater risk for being affected by the drug. And most patients - including cancer patients - experience less dramatic effects of fatigue and joint pain from the use of filgrastim - something Lyme disease patients suffer with anyway.

Scary sounding as it is to take a drug which has the risk of serious or even fatal side effects, one has to consider that if better and safer immune modulating drugs could be developed - along with antibiotics - together they might be the cure for chronic Lyme disease.

To quote from Isabel Ditrech's 2003 thesis, "Immunomodulation and new therapeutic strategies in Lyme borreliosis":

"5.3.1 Case report

A 51 year old patient with a history of frequent exposures to tick bites presented with polyarthritis in the fingers and feet. Arthritic destruction of synovial clefts mainly in the metacarpophalangial and in the proximal interphalangial joints of fingers and feet could be demonstrated by X-ray. Low, but clearly positive, serum titers of Borrelia IgG by ELISA and immunoblot (p100 +++) and a negative IgM-ELISA (both MaxPettenkofer-Institute, Munich, Germany) corroborated diagnosis of late stage Borrelia infection.  

A standard two week i.v. treatment with 2 g/day Ceftriaxone (Rocephin,Hoffmann LaRoche, Grenzach-Whylen, Germany) led to transient improvement of symptoms, i.e. subjective decline of arthritis, that lasted for eight weeks. Then, the inflammatory symptoms returned and became progressively worse, indicating that the treatment had probably failed.  

We hypothesized that persistence of Borrelia might be due to a disabled immunocompetence of the patient. Therefore, we tested whether a complete eradication of the pathogen could be achieved by combining immunosupportive treatment with antibiosis. The experimental treatment regimen, applied with the informed consent of the patient, was as follows: First week 2 g Ceftriaxone (Rocephin ) i.v. daily, second week 480 µg s.c. Filgrastim (Neupogen, Amgen, Thousand Oaks, USA) every second day, and third week 2 g Ceftriaxone daily plus 300 µg Filgrastim every second day (Figure 5.1). Neutrophil counts were determined by a Coulter STKS counter (Coulter, Krefeld, Germany)"

So this lays out the background of this individual case report on one patient. What were the results? More quoted from the above thesis:

"5.4.1 Patient case report

The combination therapy of Ceftriaxone plus Filgrastim was well tolerated. Only after the first injection of Filgrastim the patient reported acute but moderate pain in the previously affected joints i.e. the shoulder, fingers and knees. 

Circulating neutrophil counts increased from 1400 to 17000 cells/µl within 24 h after the first Filgrastim injection. Monocyte numbers increased about two-fold, while there was little effect on lymphocytes (Figure 5.2a). The plateau of neutrophil counts at about 17000 cells/µl blood was maintained until one day after the end of treatment.  

The subjective symptoms disappeared during the following six weeks after the treatment. The patient reported that he was able to resume previously abandoned sporting activities including mountain climbing and downhill skiing. Moreover, fine mechanical skills needed for piano playing were restored. 

After three months, the Borrelia IgG titer was negative. The intensity of the immunoblot at this time point was significantly reduced (from +++ to +) and two years later it was negative. Eight years after treatment the patient is still free of arthritic symptoms."

Source:
http://kops.ub.uni-konstanz.de/bitstream/handle/urn:nbn:de:bsz:352-opus-9814/Diss_formated_ENDVERSION.pdf

So it seems like at least for this patient, this method of treatment changed their life so that they could return to all the things they used to do that they loved. I would have liked to know more about this patient and how he is doing today, given it has been years since this study was completed.

And I'd like to know if a similar treatment plan would work for me and everyone else suffering with chronic Lyme disease. To take ceftriaxone and filgrastim for a couple weeks - or something similar, but with fewer side effects - only to be done with this nightmare and get on with my life would be fantastic.

It would mean no more attempts at long term antibiotic treatment and experimentation with alternative medicine. I would just get treatment for three weeks and be done with it... Sounds like a plan to me.

Reflecting on this, over the years there have been anecdotes - stories I've heard passed around Lyme disease support groups - about the occasional chronic Lyme disease patient who went on to discover they had cancer, went through chemotherapy and other supportive treatment for their cancer - only end treatment not only going into remission from cancer  - but saying that they think their chronic Lyme disease is cured, too.

These stories have been around for a while, but I've never personally known anyone who went through this process. It would be great to get a confirmation from their doctors and families that after chemotherapy and supportive treatments, they had a notable and lasting improvement and feel like their old selves again. What if a drug like filgrastim played a role in their recovery?

This isn't the only example of a chronic condition where the cause has been unknown and the symptoms can be debilitating and lead to years of loss of productivity and physical pain... let's consider chronic fatigue syndrome, also known as CFS/ME or CFSIDS.

A study completed last year in Norway showed that rituximab had a profoundly positive effect on people with CFS/ME. In this study, a few people seemed to go into complete remission from their CFS and returned to work and led normal lives. It didn't work for everyone - 40% of study participants did not experience improvement from the drug. It's unknown why. But that it worked so well for the rest of treated patients deserves a closer look because it begins to reveal the mechanisms behind what causes CFS/ME.

While there has been speculation that chronic fatigue syndrome and chronic Lyme disease (CLD) are the same condition, a recent study on the different proteins found in the cerebrospinal fluid (CSF) of both CFS and CLD patients has challenged this notion. At least in terms of objective evidence, the proteins in the CSF of both groups are different. However, what if part of the underlying process behind what causes these conditions is the same?

Quoting the above well-written article from the Phoenix Rising ME web site, let's look at the mechanism behind rituximab and what it does in people with CFS/ME:

"Rituximab is believed to deplete B-cells in two ways; by recruiting other members of the immune system to attack them and by locking on a receptor on the B-cell that tells the cell to kill itself. B-cells are an integral part of the immune response. Until they are activated, B-cells quietly troll the blood, collecting and digesting molecules called antigens that appear to be suspicious. Once they are digested they place bits of them on MHC molecules for T-cells to inspect. If the T-cells decide those molecules came from a pathogen, they turn around and turn the B-cells on – transforming them into antibody producing machines (‘plasma cells’) that can generate from 100s to thousands of antibodies per second.

These antibodies or immunoglobulins are specifically manufactured to attach to a pathogen and physically stop it from locking onto our cells. The antibodies also alert macrophages to come gobble up the pathogen and they turn on other parts of the immune system. B-cells are key players in the immune response but if they go too far; if they get too zealous, they can mistakenly attack our own cells and overactive B-cell activity has been implicated in many auto-immune disorders."

If this sounds familiar to you, then you might have been reading about Viral Genetics' targeted peptide therapy, VGV-L, for treating chronic Lyme disease.

Viral Genetics' patent states the following about treating chronic Lyme disease:

"[0116] It is believed according to the invention that Borrelia burgdorferi also produces a Toll ligand for TLR2. Replacement of the CLIP on the surface of the B cell by treatment with a thymus derived peptide with high affinity for the MHC fingerprint of a particular individual, would result in activation of the important Tregs that can in turn cause reduction in antigen-non-specific B cells. Thus treatment with thymus derived peptides could reactivate specific Tregs and dampen the pathological inflammation that is required for the chronic inflammatory condition characteristic of Lyme Disease. With the appropriate MHC analysis of the subject, a specific thymus derived peptide can be synthesized to treat that subject. Thus individuals with all different types of MHC fingerprints could effectively be treated for Lyme disease."

An easier-to-understand explanation can be found elsewhere - this research report revealed how VGV-L is used to treat HIV. In this instance, just substitute "chronic Lyme Disease" for "HIV" and you can get a picture of what VGV-L does:

"The conventional approach to HIV vaccines, for example, is to develop therapeutic vaccines to stimulate immune system response. The problem with the conventional approach is that the infected cells are camouflaged and not visible to the body’s immune system. The body’s powerful T-cells are unable to seek out and destroy the infected camouflaged cells because they cannot recognize that the cell is infected.

To understand the issue, think of the Klingon space ship on Star Trek that has its cloaking device activated. The U.S.S. Enterprise has no way of knowing where the enemy is in space. The only hope it has in winning the battle is for the Klingon vessel to be de-cloaked and, once revealed, use their ammunition to destroy it. What’s worse in the case of HIV is that while the infected cell is cloaked, it is also effectively setting off an alarm that triggers the immune system to create inflammation. Why is this important? It turns out that this inflammation is critical for allowing the HIV virus to spread to even more cells.

Many other viruses and bacteria also trigger inflammation but, unlike HIV, the inflammation does not necessarily allow or facilitate the spread of the virus or bacteria itself. However, in these cases, the inflammation itself is harmful because it creates a hostile and inflamed environment that provides the necessary components for a potential autoimmune reaction that can cause the immune system to attack and damage one’s own body. Viral believes that diseases such as Lyme Disease, Multiple Sclerosis and others involve this inflammatory mechanism.

To use the Star Trek metaphor, what Dr. Newell Rogers has developed with TPT is a de-cloaking device for the body’s immune system to use in its pursuit of invaders. Through the development and use of computational biology programs and databases, Dr. Newell Rogers and her team have created a way to remove the camouflage that is cloaking the infected cells, flagging them with custom peptides that allow the body’s immune system to seek out and destroy them.

The key discovery of the TPT platform is that a self-peptide (in other words, one that is naturally produced and a healthy part of one’s normally functioning immune system) called ―CLIP2 that was until now thought only to exist primarily inside certain immune system cells, is sometimes displayed on the outside of cells, thus leading to harmful inflammation. Dr. Newell Rogers discovered that the products of some pathogen invaders such as viruses and bacteria, when picked up on the surface of certain immune system cells, sometimes incorrectly cause those cells to display CLIP externally (i.e. ―ectopically).

Normally, when an invader strikes, this process may promote needed inflammation early in infection, but it is quickly controlled when a more specific, immune response takes over, allowing a highly-targeted immune response to be marshaled against the pathogen. However, when CLIP is improperly displayed, displayed for too long or displayed chronically, the immune system is marshaled to promote a broad and unspecified inflammation without the specific targeting, leaving open the possibility that this inflammation actually turns against one’s own cells. Replacing CLIP is the focus of Viral’s Targeted Peptides because it turns off the harmful alarm."

Read more from the source - including about individual MHC genetic profiles here: http://www.viralgenetics.com/investors/press-releases/Research_2.0_Report_Feb1_2011.pdf

They're using Star Trek metaphors to describe this... I think that's pretty geeky. Awesome.

So, it seems that whether there is current infection or not, VGV-L may be one way to effectively treat chronic Lyme disease and lower inflammation due to runaway immune dysregulation. And if infection is currently present, then it looks like VGV-L will trigger T cells that recognize the infection and summon functional B-cells to fight it.

Now, getting back to Isabel... Remember Isabel, the researcher who used filgrastim and ceftriaxone to treat a patient with chronic Lyme disease about a decade ago? Yes, that Isabel.

Well, she wrote another paper, along with Rauter, Kirshning, and Hartung: "Borrelia burgdorferi-Induced Tolerance as a Model of Persistence via Immunosuppression"

The abstract states:

"If left untreated, infection with Borrelia burgdorferi sensu lato may lead to chronic Lyme borreliosis. It is still unknown how this pathogen manages to persist in the host in the presence of competent immune cells. It was recently reported that Borrelia suppresses the host's immune response, thus perhaps preventing the elimination of the pathogen (I. Diterich, L. Härter, D. Hassler, A. Wendel, and T. Hartung, Infect. Immun. 69:687-694, 2001). Here, we further characterize Borrelia-induced immunomodulation in order to develop a model of this anergy. 

We observed that the different Borrelia preparations that we tested, i.e., live, heat-inactivated, and sonicated Borrelia, could desensitize human blood monocytes, as shown by attenuated cytokine release upon restimulation with any of the different preparations. Next, we investigated whether these Borrelia-specific stimuli render monocytes tolerant, i.e. hyporesponsive, towards another Toll-like receptor 2 (TLR2) agonist, such as lipoteichoic acid from gram-positive bacteria, or towards the TLR4 agonist lipopolysaccharide. Cross-tolerance towards all tested stimuli was induced. Furthermore, using primary bone marrow cells from TLR2-deficient mice and from mice with a nonfunctional TLR4 (strain C3H/HeJ), we demonstrated that the TLR2 was required for tolerance induction by Borrelia, and using neutralizing antibodies, we identified interleukin-10 as the key mediator involved."

Source: http://iai.asm.org/content/71/7/3979.full

Where have I heard something like this before? Oh, Dr. Karen Newell Rogers - that's right - she discussed this at a recent Lyme disease research conference:

"[...]Some researchers would argue that chronic inflammation requires the continuous presence of bacteria, whereas others would suggest that continuous presence of bacteria does not always result in inflammation and that exacerbations of chronic symptoms could result from infection with a different organism--or that chronic symptoms could re-cur from unrelated pro-inflammatory events. Potentially reconciling these seemingly conflicting perspectives on the mechanism of Lyme disease may be the effect of Borrelia burgdoreri’s bacterial by-products on Toll Like Receptors, (TLR)-mediated immune activation. 

TLR appear to be the “gate-keepers” of an inflammatory response. Bacteria, including Borrelia, produce products that, by binding to TLRs on the cell surface, promote leukocyte activation, cytokine production, and acute inflammation. In some genetic backgrounds of mice, acute inflammation is sufficient to fight off infection and resolve disease. In other mouse strains, the pathogens, or in this case the bacteria, get past TLR-induced inflammation and remain symptomatically undetectable in cells and tissues (Barthold, etc); Barthold et al. have found that no matter how severe or mild the disease in any of the genetically inbred strains of mice, there was no more inflammatory disease when the bacteria were eliminated."

And where else have I heard about IL-10 production before? Oh, right - Rituximab, and research on gender differences in antibody response to Borrelia burgdorferi...

From the previously mentioned Phoenix Rising ME article:

"While Rituximab is busy destroying B-cells there is also evidence that it may actually be turning on NK cells – which, of course, habitually underperform in CFS. Rituximab also appears to increase production of IL-10 – a key anti-inflammatory cytokine that may be a protective agent in ME/CFS – and reduces levels of the powerful pro-inflammatory cytokine tumor necrosis factor. A review article suggested that Rituximab was able restore Th1/Th2 balance in the immune system. These results suggest Rituximab could be working as an immunodulator helping to re-balance the immune response by turning down the over-activated parts of it and bumping up the under-active ones."

All this ties together quite nicely, it seems, with other research I have listed here - forming a master hypothesis with different pieces. Does it hold up to scrutiny? Tell me - I'd love to hear your ideas.

But here is the master hypothesis, in its infancy:

1) Host genetics play a role in the ability of mice (and possibly people!) in clearing Borrelia burgdorferi infections. See:

http://campother.blogspot.com/2011/08/immune-infection-hla-dr-alleles.html

The host's genetic background in developing chronic infection is, however, open to debate - and may not play as big a role in disease as Borrelia burgdorferi s.l.'s genetic diversity/VlsE recombination on different plasmids:

http://campother.blogspot.com/2011/08/do-different-genetic-haplotypes-matter.html
http://campother.blogspot.com/2011/08/more-on-genetic-haplotypes-and-lyme.html

2) The genetics of Borrelia burgdorferi strains play a role in how quickly they disseminate into host tissues and also how well they can generate inflammation - which leads to overstimulation of the immune system in production of poor quality plasma b-cells, but also, ironically, immune suppression because of the mechanisms Isabel Diterich and Karen Newell Rogers describe. Refer, also, to Tunev and Barthold et al's research, "Lymphoadenopathy during Lyme Borreliosis Is Caused by Spirochete Migration-Induced Specific B Cell Activation":

http://campother.blogspot.com/2011/06/paper-borrelia-burgdorferi-rst1-ospc.html
http://spirochetesunwound.blogspot.com/2011/07/does-borrelia-burgdorferi-cause.html
http://www.plospathogens.org/article/info%3Adoi%2F10.1371%2Fjournal.ppat.1002066
http://spirochetesunwound.blogspot.com/2010/07/antigen-presentation-in-bloodstream-how.html (refer to other research on relationship between b-cells/plasma cells and T cells)

It could also be that not having enough iNKT cells is an issue:
http://www.pnas.org/content/105/50/19863.full.pdf

2a) The changing pattern of antigenic variation during this time may also be why patients produce an undulating immune response in measured antibodies which echo a more drawn-out response similar to relapsing fever:

http://campother.blogspot.com/2012/02/paper-course-of-antibody-response-in.html
http://campother.blogspot.com/2011/08/antibodies-linked-to-long-term-lyme.html (read comments, too)
http://www.ncbi.nlm.nih.gov/pubmed/9108482
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2772371/
http://www.ncbi.nlm.nih.gov/pubmed/11544329
http://campother.blogspot.com/2011/07/lyme-disease-western-blots-and-antigen.html

It may not be that the tests are lousy for measuring antibodies which are present to Borrelia burgdorferi. It may be that the antibodies are not present because they are tied up in immune complexes.

2b) There is also the possibility that Borrelia burgdorferi is occasionally intracellular in nature, though there is not enough in vivo evidence to support this. If so, it would also explain why an undulatory immune response might be present:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3067508/?tool=pubmed
http://campother.blogspot.com/2011/07/fibroblasts-and-lyme-disease-sample.html

Whether or not items #2a and #2b are relevant here remains to be seen - the main point is that Borrelia burgdorferi can lead to both overstimulation of the immune system as well as immune suppression.

Based on this, I surmise that may not be that blood tests are so lousy at detecting antibodies produced by the presence Borrelia burgdorferi. It may be that there is no reliable way to detect the presence of infection by correlating them with the presence of antibody responses (seronegative Lyme disease).

3) What gender you are and your hormone levels and metabolism may play a role in persisting symptoms and prolonged infection as well, so there is ALSO a metabolic cause behind chronic Lyme disease. How well the immune system can respond to initial infection to begin with seems to play a role in developing chronic Lyme disease, as even 10% of acute cases of Lyme disease result in treatment failure.

http://campother.blogspot.com/2012/03/lyme-disease-presents-differently-in.html
http://campother.blogspot.com/2012/01/two-new-hypotheses-for-chronic-lyme.html (read comments, too)
http://www.ncbi.nlm.nih.gov/pubmed/17438273 (this may provide the scientific link for the anecdotes that people who develop chronic Lyme disease generally were under more stress when they contracted the disease)

4) If there are persister cells, this is an additional consideration - throwing more antibiotics at a pathogen which is antibiotic tolerant when it is a persister cell will, at most, keep the infection from getting worse but it won't eliminate it.

http://campother.blogspot.com/2012/01/paper-persistence-of-borrelia.html
http://campother.blogspot.com/2012/02/blog-log-spirochetes-unwound-on.html

See also:
The research of Kim Lewis on persister cells: www.bu.edu/abl/files/killing_persisters.pdf
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3145328/

And it may be that persister cells are more likely to be on the scene earlier, depending on how appropriate a given antibiotic is for treating specific genospecies - refer to item #2 above, but also:

http://campother.blogspot.com/2011/05/abstract-evaluation-of-in-vitro.html

5) Because the host has a sub-optimal immune system, even with long term antibiotics, a subset of the population will have trouble clearing the remaining spirochetes after antibiotics are stopped. Additional antibiotics plus a treatment which eliminates low quality plasma b-cells and promotes the activity of Treg cells which recognize current infection could overturn the dysregulated immune system.

What does this boil down to?

Easy: The argument of "is it a chronic infection or is it an immune disorder, possibly autoimmune" is a false dichotomy and too simplistic.

The circumstances which give rise to chronic Lyme disease are more complex than that, and if people want to solve the chronic Lyme problem, they have to roll up their sleeves and look at more puzzle pieces and how they fit together.

Image credit: 
Original image by Muns on Wikimedia Commons; derived image above by Schlurcher.

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

0 News: Judy Mikovits To Be Released On Bail

The latest report on the Mikovits case comes from the AAAS Science Insider:
http://news.sciencemag.org/scienceinsider/2011/11/inmate-mikovits-meets-judge.html?ref=ra

Regarding the outcome of today's proceedings, the judge granted a request for a continuance on the extradition demand and asked Mikovits to return on the 19th of December.

Mikovits, who has denied wrongdoing through lawyers handling her civil suit, posted bail shortly after the hearing and has been expected to be released this evening.

More details on the case and discussion of a settlement can be found here, on the Ventura County Star site:
http://www.vcstar.com/news/2011/nov/22/world-known-researcher-set-to-be-released-on/

And here, Nature blog discloses that one of Mikovits' fellow researchers, Max Pfost, claims to have stolen her notebooks and to have given them to Mikovits', but Mikovits' denies this.

Nature blog also states:

The affidavits add a new wrinkle, alledging that Mikovits, a chronic fatigue syndrome researcher, was scheming with Pfost to “move the grants from WPI”. This includes about half of a US$1.5 million research grant (an R01) from the National Institute for Allergy and Infectious Diseases. In a meeting at a bar the night Mikovits was fired, Pfost claims: “She stated she was going to try to move the RO 1 grant and the Department of Defense (DOD) grants and stop the Lipkin study.”

More here: http://blogs.nature.com/news/2011/11/researcher_confesses_to_steali_1.html

The plot thickens... What really happened has yet to be seen.

Read More

0 Article: The Mouse Trap: How One Rodent Rules The Lab

Slate recently posted a three part series on the use of mice in biomedical research and how that use has helped as well as hindered drug development and scientific understanding of human illness.

This three part series kicks off with "The Mouse Trap: the dangers of using one lab animal to study every disease", which examines the rise in the use of the murine (mouse) model  to test new pharmaceuticals and get a better understanding of human processes in illness. We also learn that this model - useful as it has been - has its drawbacks.

 Mark Mattson, neuroscientist, finds that murine studies may not accurately represent what happens between the study and control group when the control group is often overfed, sedentary, and develops cancer, renal problems, and diabetes - unlike a wild mouse living in its normal habitat. It was later discovered that medication which worked in chubby rodents did not go on to work in rodents with a normal weight and activity level.

Furthermore, in clinical trials on humans later on, the difference between plump mice and a diverse group of humans was too great. A drug which worked in the rodents did not work in humans at all.

Cheap, efficient, and mass produced, these mice are raised in germ-free barrier rooms and sometimes pumped with antibiotics. The ease in which they can be raised, bred, and genetically modified to single out specific processes similar to those in the human body to study has made mice very popular for researchers. While they can closely model many processes in the human body, are they a close enough match for modeling and testing everything?

Mattson argues they are not, and that science needs to wake up and take a closer look at how heavily mice have been used to make decisions that affect human medicine. He's not the only one, either - Clif Barry, the nation's leading expert on tuberculosis, thinks the murine model has shortcomings too. This is no clearer than in work in his own field: The latent form of TB that humans get is not one which can be found in mice. Mice do not experience a latent stage of TB - they only get progressively weaker, age, and die.

This particular passage from the article gave me a moment to pause and reflect:

"[...] for some patients a latent case of tuberculosis can suddenly become active. The granulomas rupture and propagate, spilling thousands of organisms into the lungs, where they can be aerosolized, coughed up, and passed on to a new host. Left untreated, the infection migrates into the bloodstream and other organs; widespread inflammation leads to burst arteries or a ruptured esophagus; and in about half of all cases, the patient dies.

The layered granuloma is the defining feature of human tuberculosis: The place where the host fights the infection (successfully or not), and the necessary site of action for any drug. To cure the disease, a treatment must be able to penetrate each ball of cells, whatever its type or composition; every last bacterium must be destroyed. "It's the structure of those granulomas that makes it so difficult to treat TB," says Barry. And they simply don't exist in mice.

If you infect a mouse with TB—if you spritz a puff of infected air into its nostrils through a trumpet, as so many labs do around the world—the animal's lungs quickly fill up with bacteria and immune cells, like a nasty case of pneumonia. There are no discrete balls of tissue, no well-defined granulomas sheathed in fibrin, no array of structures that harbor the bugs at various stages of development. The mice have no special, latent form of TB, either, and no way to pass on the disease. They simply die, after a year or two, of a slow and progressive decline.

That's why we've made so little progress using mice to generate new drugs and treatments, Barry tells me."

The article continues to outline the history behind why mice have become the dominant research model to use to determine whether or not a drug or treatment plan will be tried in clinical trials on people, and also discusses the setbacks generated by this decision. In the end, the verdict is that using mice has distinct limitations and a number of studies may be invalidated and useless because of it.

It does get me to wondering, of course, as to how useful the murine model is for studying Borrelia burgdorferi. Perhaps in some ways it is useful, but in others it is less so: Dr. Stephen Barthold has said mice are a poor model for neuroborreliosis.

And this does raise the question of which details are important in a study that the layperson will miss when reading it. The average person will read about the study design, and perhaps some discussion and the conclusion - but will one question the decision to use one research animal model over another? What about primers? What about media? What about any number of small details that once changed could result in a different outcome?

Read the full Slate article, and continue with the next two articles in the series. It is a fascinating look into the world of biomedical research, tradition, and profit.

Reference: http://www.slate.com/articles/health_and_science/the_mouse_trap/2011/11/lab_mice_are_they_limiting_our_understanding_of_human_disease_.html

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

1 Chalcolithic Iceman Had Lyme Disease Over 5,000 Years Ago

Ötzi, The Iceman, Cometh...

Thanks goes out to Joanne, from the Looking At Lyme Disease blog for posting about an article from National Geographic on the Iceman, a mummified man who had died over 5,000 years ago and has been a treasure trove of information for scientists ever since.

The original article is found here:

Iceman Autopsy - Unfrozen: There was only one way scientists could unlock the mystery of the famous Iceman. Take away his ice.


Source Link: http://ngm.nationalgeographic.com/2011/11/iceman-autopsy/hall-text

The entire article outlines the process of extracting and examining tissues and food from within the mummy - an interesting read in itself. But the part that will stand out for Lyme disease patients is this bit:

"Perhaps most surprising, researchers found the genetic footprint of bacteria known as Borrelia burgdorferi in his DNA—making the Iceman the earliest known human infected by the bug that causes Lyme disease."

It would be interesting to know exactly which Borrelia burgdorferi strain was found and in which tissue sample(s) it was found.

And reflecting on this, this indicates that Borrelia burgdorferi has been around for a long time and in Europe for a long time.

More Information On The Iceman:
http://en.wikipedia.org/wiki/%C3%96tzi_the_Iceman

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

0 How To Avoid Sample Contamination

A quick and dirty primer for those who want to ensure that their samples do not get cross-contaminated with DNA from other bacteria, viruses, etc.:

1. Use aerosol barrier pipette tips.
   
2. UV-irradiate all workstations used for the setup of master mix preps and PCRs.
   
3. Treat all surfaces and tube racks with a 10% bleach solution.
   
4. Use frequent and careful glove changes.
   
5. Perform DNA extraction, PCR setup, and PCR product analysis in different rooms.
   
6. Use clean systems.
7. Use a negative control such as UV-treated, deionized water.
8. Do not do bacterial work, etc. during any human DNA extraction.

Follow these simple tips, and then when others look at your research and claim that Borrelia burgdorferi could only be found where it was due to  sample contamination, point out that a number of steps were taken to prevent it.

At what point when an experiment is repeatedly reproducible does one stop saying the end result was due to sample contamination and begin saying the end result was genuine?

What other steps can you take to prevent and eliminate sample contamination?

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

1 Paper: Evaluating Research Quality

This is a brilliant, well-written publication, Evaluating Research Quality, on how to assess the quality of research you are reading - written by Todd Litman at the Victoria Transport Policy Institute - which is "an independent research organization dedicated to developing innovative and practical solutions to transportation problems".

Read Evaluating Research Quality Here: http://www.vtpi.org/resqual.pdf

Todd provides a number of examples of what makes good, well-informed research and what is biased, ill-informed research, drawing from examples in the transit sector but also general examples such as the source of statements such as "Eskimos (Inuit) have 23 words they use to describe snow" and ideas such as "AIDs doesn't kill people, antiviral medication kills people".

The entire document is well worth a look from start to finish, and the "Sixty Four Methodological Potholes" table is an interesting and extensive read. It's more detailed than Carl Sagan's Baloney Detection Kit and highlights some of the same illogical fallacies and bias some have presented in their publications and arguments.

This specific portion could be applied (and I should probably more stringently apply these pointers to papers reviewed here) to Lyme disease research and all research in general, so I want to share it. Credit goes to Todd Litman and the Victoria Transport Policy Institute, who allow republication with credit. (Thanks, guys.)

Research Document Evaluation Guidelines

The guidelines below are intended to help evaluate the quality of research reports and articles.

Desirable Practices

1. Attempts to fairly present all perspectives.
2. Provides context information suitable for the intended audience. This can be done with a
   literature review that summarizes current knowledge, or by referencing relevant
   documents or websites that offer a comprehensive and balanced overview.
3. Carefully defines research questions and their links to broader issues.
4. Provides data and analysis in a format that can be accessed and replicated by others.
   Quantitative data should be presented in tables and graphs, and available in database or
   spreadsheet form on request.
5. Discusses critical assumptions made in the analysis, such as why a particular data set or
   analysis method is used or rejected. Indicates how results change with different data and
   analysis.Identifies contrary findings.
6. Presents results in ways that highlight critical findings. Graphs and examples are
   particularly helpful for this.
7. Discusses the logical links between research results, conclusions and implications.
   Discusses alternative interpretations, including those with which the researcher disagrees.
8. Describes analysis limitations and cautions. Does not exaggerate implications.
9. Is respectful to people with other perspectives.
10. Provides adequate references.
11. Indicates funding sources, particularly any that may benefit from research results.

Undesirable Practices

1. Issues are defined in ideological terms. “Straw men” reflecting exaggerated or extreme perspectives are use to characterize a debate.
2. Research questions are designed to reach a particular conclusion.
3. Alternative perspectives or contrary findings are ignored or suppressed.
4. Data and analysis methods are biased.
5. Conclusions are based on faulty logic.
6. Limitations of analysis are ignored and the implications of results are exaggerated.
7. Key data and analysis details are unavailable for review by others.
8. Researchers are unqualified and unfamiliar with specialized issues.
9. People with differing perspectives are insulted and ridiculed
10. Citations are primarily from special interest groups or or popular media, rather than from peer reviewed professional and academic organizations.

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

0 Abstract: Interaction of Borrelia burgdorferi in coculture with human fibroblasts

I haven't seen this paper mentioned elsewhere - this is from a conference held last year. Need to find full text - not available on ASM or Scholar...

Interaction of Borrelia burgdorferi in coculture with human fibroblasts 
Daniel Wilfinger, Gerd Leitinger, Anna Maria Pabst, Helmut Schaider, Elisabeth Aberer

Kurzfassung/Background: B. burgdorferi (B.b.) can be recovered long after initial infection from antibiotic-treated patients; a protective effect of fibroblasts was assumed. Outcome of previous studies differ whether spirochetes are able to invade fibroblasts. Skin fibrosis has repeatedly been observed in European chronic Lyme borreliosis. So the aim of our study was to examine the interaction of B.b. with fibroblasts in coculture ultrastructually and to investigate key factors for fibrosis such as transforming growth factor ß (TGF-ß) and the production of type I collagen.

Materials and Methods: Human skin fibroblasts were propagated in Dulbeccos’s modified medium at 37°C and 5% CO2 up to 10 5 cells per culture. Bb sensu stricto and B. afzelii were cultured at 34°C in BSK-H medium up to 10 8 cells per culture. Borreliae and fibroblasts were then coincubated in RPMI medium at 37°C for 14 days. Cocultures and fibroblasts only were harvested between 2 and 14 days 3-4 times, fixed in glutaraldehyde and prepared for electon microscopy analysis. Supernatants were investigated for TGF-ß by ELISA. mRNA levels for type I collagen were measured by real time PCR.

Results: Electron micrographs showed borreliae which were able to attach to the fibroblast membrane through protein bridges. Single spirochetes seemed to pervade fibroblast cytoplasm by invagination surrounded by an intact fibroblast membrane. Structural changes of extracellular borreliae due to the adverse culture condition were observed. Shedding of outer membrane blebs forming granules and tubules were seen as well as cystic degeneration of borreliae.

After 2 days of coculture with B.b.s.s. the total amount of TGF-ß in the supernatants was approximately the same as in fibroblast-cultures, after 14 days of coculture production of TGF-ß was decreasing in cocultures with B.b.s.s.. In supernatants of cocultures with B. afzelii TGF-ß was not detectable at any time.

The mRNA-levels for type I collagen were elevated after 2 days of coculture with B.b.s.s. as well as in cocultures with B. afzelii in comparison to fibroblast-cultures. After 7 days the mRNA-levels for type I collagen were decreasing in cocultures in comparison to fibroblast-cultures, a stronger decrease was observed in cocultures with B. afzelii.

Conclusions: The interaction of B.b.s.s. and B. afzelii with human fibroblasts was verified by electron microscopy. Fibroblast integrity was not disturbed by borreliae. Intracellular accumulation of spirochetes was not detectable.

We demonstrated differences in the two species B.b.s.s. and B. afzelii concerning production of type I collagen and TGF-ß. In contrast to cocultures with B.b.s.s. TGF-ß was not detectable in cocultures with B. afzelii at any time. In line with these findings in cocultures with B. afzelii a stronger decrease of collagen production was observed in comparison to cocultures with B.b.s.s. after 7 days of coculture. The elevation of mRNA-levels for type I collagen in cocultures after 2 days is confirmed by literature: Scleroderma-fibroblasts also show an increase in collagen production after 2 days of culture.

Schlagwörter/Keywords : Borrelia, B. burgdorferi sensu stricto, B. afzelii, electronmicroscopy, morphology, TGF-ß, mRNA collagen, morphea, fibrosis, ACA

Read More

0 BBC Article On Assessing Risk (Cellphones, Carcinogens, & Others)

This isn't about Lyme disease and tickborne infections, but it is definitely food for thought when considering the risk involved in any action you decide to take.

Check this article out:

Link: http://www.bbc.co.uk/news/magazine-13886254

Summary Teaser:

BBC News - Go Figure: Do we understand 'risk' of mobile phone use?

What should we make of recent news reports speculating about whether mobile phones cause cancer? It's all about how we deal with uncertainty, says Michael Blastland in his regular Go Figure column.

I highly recommend reading the highly rated comments from the "All Comments" pile, too. Insightful stuff, there.

Read More

0 Blog Log: My Lyme Disease Story

I wanted to point out a relatively new Lyme disease blog on the block, My Lyme Disease Story.

The author has been a practicing MD who was bitten by a tick somewhere inside of Yosemite in 2010, developed the EM rash and other symptoms, was undertreated, and eventually sought the care of an LLMD.

I usually don't write much about personal blogs, tending to focus on science-oriented ones instead - but this is a rare occasion to see how a doctor approaches the same conditions and illnesses their patients must face. So far, she is facing Lyme disease with blunt honesty and eloquence - her recent description of suicidal ideation is a well-written post on the darker nights and thoughts of many living with pain and loss of functionality.

This most recent entry is a hard read and not for the faint of heart - it may prove triggery for some:

http://mylymediseasestory.blogspot.com/2011/06/depths-of-darkness-and-despair.html

I recommend not stopping there, and reading all entries to date.

And if you are reading this and are a Lyme disease/tickborne illness patient, please stop by and offer words of support and encouragement - as well as your knowledge and experience. This is the beginning for her and I hope she does well and recovers soon.

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

2 Media Watch: Lyme Disease in Huffington Post & Baltimore Sun

CC Photo Credit: debwire

The past couple of days have seen the publication of two opinion pieces on Lyme disease online: Dan Rodricks' commentary, "MPT airing deeply flawed film about Lyme disease", published June 19 in The Baltimore Sun (121 comments and counting), and Leo Galland, MD's, "Lyme Disease Symptoms:Key Facts About This Mysterious Illness", published June 18 in The Huffington Post (278 comments and counting).

The first article is an op-ed piece about how the film, Under Our Skin, was pulled off the air and not shown by one PBS affiliate station in Boston - but is being shown on Maryland's PBS stations. Mr. Rodrick writes about how Philip Baker, research scientist at the NIH and executive director of the ALDF, views the film as being dangerous to public health and "a one-sided and emotionally charged attack on science". The position maintained by Mr. Rodrick is that after the 2009 IDSA Lyme disease panel guidelines review (part of an investigation conducted by Attorney General Blumenthal) and the 2010 report on that review, the gig is up - science has determined that the standard treatment guidelines are proper for the treatment of Lyme disease.

The second article is one doctor's perspective on which symptoms can be present with Lyme disease and what he states are facts about the disease - while making some jabs at the IDSA. Dr. Galland's position is clear: he believes that Lyme disease can be a persistent infection that survives standard antibiotic treatment and that seronegative Lyme disease happens (how often is not stated here either way).

Figures stated in Dr. Galland's short article conflict with Mr. Rodrick's - Dr. Galland states that over 100,000 people contract Lyme disease each year; Mr. Rodrick says it's 30,000 people and fewer people than contract chicken pox annually - as if that is somehow a valid or even worthwhile comparison to make.

They are both wrong, of course: The total number of confirmed cases of Lyme disease that were reported to the CDC in 2009 is 30,000, plus there were fewer than 10,000 probable cases reported that year - but no one knows exactly how many people have been infected.

Even the CDC has stated:

"Between 2008 and 2009 there was a 3.6% increase in confirmed cases and 35.6% increase in probable cases. Much of the increase can be attributed to variability in surveillance practices, although evidence of true emergence exists in certain areas. Because of the burden on endemic states posed by Lyme disease surveillance, some states have modified surveillance protocols to better manage limited resources. States using modified methods, including case estimation, might report decreased case counts."

So a true, hard number is probably never going to be knowable for a variety of reasons, and whatever estimate you supply depends on who you ask and how the data were collected - ask Wolfram Alpha and others where they got their Lyme disease data, how they weighed it, and where extrapolations were made and how. Ask someone else. I'm sure the answers will be different, and again - is there any way to ensure a more accurate estimate if epidemiological studies are cut short due to lack of funding?

Dr. Galland states the IDSA says 95% of Lyme disease patients are cured and that he finds even just a 5% failure rate unreasonable for a bacterial infection (we are all really concerned about MRSA's failure rate, to be certain) but even within the IDSA's own 2006 Lyme disease treatment guidelines, it is stated that there is an up to 10% treatment failure rate in early infection.

Taken from those guidelines:

"Less than 10% of individuals do not respond to antibiotic therapy, as evidenced by the presence of objective clinical manifestations, and rarely is re-treatment required. In general, patients who are more systemically ill (e.g., febrile with significant constitutional complaints) at the time of diagnosis take longer to have a complete response to therapy."

That's early infection - and not the controversial chronic Lyme disease that has been subject to debate. This alone should be concerning to anyone who is bitten by a tick and is infected.

In terms of the responses to each of these articles, I shouldn't be surprised: once again, the Wall Of Polarization has the opportunity to rear its ugly head in comments. This brings me no small amount of grief to read some of the responses (which I am still weeding through and trying to catch up on, knowing full well by posting this entry that more comments will be added to the fray).

My problem with all of this is that both views tend to be oversimplifications which are viewed as suitable grist for the ratings mill - but neither of these articles truly dig into microbiology (and molecular biology) to show how one could match up the rationale for supporting certain treatments (e.g. longer courses of antibiotics) or denying their effectiveness for what may be happening in the host in vivo.

Nowhere in any of these online articles do I see mention of discussions with content such as this:

Tuberculosis is treated longer term with a combination of antibiotics in part because it is intracellular and it can persist. In several studies, there is evidence that mostly extracellular Borrelia burgdorferi is at least conditionally intracellular - but when, and how often? Do we know - or is this something which requires more research, noting here that intracellular obligate parasites are difficult to study? This is important, because their intracellular nature - or lack thereof - would change the duration and form of antibiotic treatment.

Can Lyme disease bacteria persist in the host after antibiotic treatment? Yes, they can persist. But what are they doing there? Are they infectious and can they replicate? Some researchers say yes; some say no. Given this state of affairs, more research on pathogenesis needs to be done.

Can some genotypes result in more severe symptoms of infection? Yes, they can - and if this is so, is it possible that different genotypes require somewhat different (and possibly extended) treatment?

But these questions are not raised in news articles and op-ed pieces online. Relying on expert opinion often is - without any investigation of the foundation supporting that opinion. And this is where these articles fall short for me, in both cases - here an opportunity to engage the public in science education is squandered.

Presented with two opposing points of view, the casual observer is left with a sense of confusion after reading both sides. One makes their appeal to reason and the IDSA's expertise; the other makes their appeal to reason and their clinical expertise; the patient makes an appeal on behalf of their own human suffering. It is not too uncommon to see others respond to this polarization by throwing their hands up in disgust and walking away, not knowing what the truth of the matter is - but in the end, I hope that somehow the gap can be bridged between the search for scientific fact and tending to the chronically ill with care and respect.

At the end of Mr. Rodrick's commentary, he leaves a quote by Philip Baker:

"I believe in providing equal time to those with evidence to support their claims," says Mr. Baker. "But in this case, there is none. One may have differing opinions on matters related to religion, philosophy and politics. However, when it comes to science, peer-reviewed evidence is the gold standard. The producers and advocates of this slick film have no such proof that merits public viewing."

I both disagree and agree with this quote. I think there already is evidence to support some of the claims of those who think Lyme disease can be a persistent infection, and that more research in this area is necessary. I also think that while Under Our Skin has been a great film for portraying the symptoms and trials which patients must endure, it had very limited discussion of the pathogenesis of Lyme disease Borrelia and discussion of scientific research that supports evidence for persistent infection as well as other causes for persisting symptoms.

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More

1 I've Got You... Under My Skin

There you folks go.

Seriously, if you stopped for just one moment and responded to the naysayer's comments with evidence, then the conversation might be more productive...

Exchange online, identities removed and exact words modified because they aren't necessary to make the point - and besides, one was anonymous, anyway:

Party A: 

"I was not impressed by this movie. It is a rant against Big everything. There's no explanation of what is wrong with the guidelines. I'd like to see LLMDs explain what chronic Lyme disease is, explain which guidelines are in error and why, and then present research which contradicts research cited in the guidelines. I don't expect this will happen any time soon. I don't understand why everyone is so conspiracy theory about this and isn't looking at the problem logically.

Why would Big Pharma not want use of long term antibiotics since it's money making?

Further Comments from (assumed same) Party A:

"The guidelines have research citations. I have no issue with questioning them, it's how better ones are made. But any change must be based on research not anecdote. When someone can define what "persistent Lyme disease is" and use DB RCS with the right methodology and show longer term antibiotic therapy is good then the guidelines will change."

(Remainder of comment scrapped for repeating much of previous comment made above.)

Party B response to Party A:

"Clearly, you are clueless - even after watching the film. If you ever get Lyme disease (chances are high, considering it's an epidemic), you won't like these guidelines so much."

Party C response to Party A:

"You sound just like these two guidelines supporters and authors of guidelines. If you want to know more about it then read. Or better yet get bit by a tick and get sick and see what life is like for a Lyme disease patient."

(Remainder of comment thread removed for brevity.)

Hi, I'm sorry you are suffering. This disease sucks, and I hate it, too.

Unfortunately, this isn't making a strong case in support of the issues we're facing here and providing a well reasoned counterpoint to Party A - whoever they are - is important. (And they could be anyone who is skeptical and wants to know the answers to their questions.)

If you can convince them of the problems patients are facing, then you can convince just about anyone.

I suggest that you read the following links:
http://campother.blogspot.com/2011/05/chart-ways-to-discuss-your-position-in.html
http://campother.blogspot.com/2011/05/repost-lyme-disease-rant-wall-of.html

You can win in an argument - or at least present information which will get others to think, including those observing your exchange who do not comment at all.

Remember that for every comment you write and receive, there are hundreds of people who say nothing and your comment and your post has an effect on them.

This work by Camp Other is licensed under a Creative Commons
Attribution-NonCommercial-ShareAlike 3.0 Unported License.

Read More