Date: October 19, 1999
Interstitial Cystitis Network - Chat Log (www.ic-network.com)
Topic: Pediatric Urology in Collagens
Speaker: Pamela Howard, University of Pennsylvania, Philadelphia

<icnmgrjill> Welcome to the ICN "Meet the Expert Chat" for October 19, 1999. Our guest speaker is Dr. Pamela Howard. One of the pleasures of welcoming IC researchers to the ICN is that we all get to learn just a bit more about the complex issues that researchers face. While we, the IC patient, want a simple treatment that will work TODAY! Researchers and physicians have to focus on the details of anatomy the details of how the bladder works and the details of how medications interact with the body. In tonight's case, we will have the opportunity to learn just a bit more about how the bladder membrane is structured and how that structure can change. We all know that there are some patients who develop smaller, stiffer bladder walls that holds less urine. Dr. Howard is at the forefront of research that explains how this could be happening. This is very exciting!

<icnmgrjill> Dr. Howard, welcome!

<pam> Thanks for the invitation! I'm very pleased to be here and I hope that I can add some information to help any or all of you who are here.

<icnmgrjill> We have a presentation tonight that is fascinating! Let's go ahead and then we'll take your questions from the floor. And now... welcome to the world of an IC researcher!

--------------PRESENTATION BEGINS----------------

Thank you for the invitation to discuss my research on pediatric bladder tissue and cells, and how this may relate to interstitial cystitis. The title of the published study using the pediatric tissue is "Molecular Analysis of Collagens in Bladder Fibrosis." I would just like to give a little background concerning the relevance of *collagen* and *fibrosis* in the bladder, describe the patient population from which the tissue samples were derived, and then discuss how this may relate to interstitial cystitis.

Collagens are a family of proteins which form the structural framework of almost all organs. This family consists of more than 20 different collagen types, each type is classified by Roman numerals, i.e., type I, type III, etc. Some collagen types are restricted to certain parts of the body (such as type II collagen which is found in cartilage), while other types are found ubiquitously (such as type I collagen).

Even though there are many types of collagen, all collagen molecules share similar important structural features. All collagen molecules consist of 3 polypeptide chains (called alpha (a) chains). These chains wind around one another and form a triple helix, like the strands of a rope. The collagen molecules associate with other collagen molecules (the same type or a different type) to form fibrils, and then larger fibers, and the fibers are additionally strengthened by chemical bonds. This composition of collagen fibers is important since the structural integrity of a tissue or organ is based on the three-dimensional organization of the collagen molecules.

In general, collagens are noted for their very high tensile strength, lending support to an organ and a resistance to pulling forces. This is particularily true for the *fibrillar* collagens which form cable like fibers. Types I, II, III and V are examples of fibrillar collagens and type I and III collagens are the major collagens found in the bladder. Other collagens do not form fibers, per se, but exist as a meshlike sheet which serves to separate different structures. An example of this is type IV collagen, which is a component of basement membranes which forms a sheet like structure at the basal aspect of the urothelium in the bladder and serves to separate the urothelial cells from the underlying connective tissue. Type IV collagen also surrounds bladder smooth muscle cells in a cocoon-like fashion. It also is present at the interface of Schwann cell - axonal processes in the peripheral nervous system and its presence is necessary for myelination to occur.

The most abundant collagens in the body, and in most organs, are types I and III collagen. These collagens form a network of structural fibers that connect the cells of the organ and also connect other structural components. The specific types of collagens, the amount of these collagens and how they are arranged in tissues and organs depends upon the specific organ and its function. If there are changes in the amounts of these collagens, in the ratio of one type to another in the fibers, and/or in the normal arrangement of the fibers, then there can be a pathological change in function.

In the bladder, types I and III collagen make up the majority of the collagenous proteins in that organ. Overall in the bladder wall, approximately 75% of the collagen is type I and 25% is type III. These collagens are found as a network of fibers in the layer of the bladder beneath the urothelial lining cells, and also around the smooth muscle bundles in the outer layer of the bladder wall. During the filling phase of the bladder, the collagen fibers actually shift from their A relaxed* orientation and expand (just like a coiled telephone cord when it is pulled). The collagen between the relaxed muscle bundles also extends. This causes the bladder to fill normally without any increase in pressure. It is thought that, at the limit of this extension, the pressure rises, the muscle cells then contract, and the bladder empties. So, normally, the collagen fibers in the bladder help (1) to maintain the tissue architecture and strengthen the bladder wall, (2) allow the bladder to expand without any pressure increase, but at the same time, (3) limit the expansion so that micturation can occur within a normal volume and pressure range.

Fibrosis is characterized by an excessive or abnormal accumulation of collagens in the tissues and organs which results in a destruction of the normal tissue architecture and compromised function. Fibrosis could also be defined as abnormal wound healing. Normally there is a closely regulated sequence of events, a defined process of repairing an injury, termed wound healing. The injury could result from one of many sources. Examples of this are trauma, infection, chemical injury, mechanical injury, inflammation, diabetes, hypertension, and autoimmunity, to name a few. The injury causes cells within the organ to become activated and increase their output of collagen, and also, possibly decrease its degradation. The end result is an increase in the amount of fibrous collagens. Normally, once the injury is repaired, the process is turned off. In fibrosis, the process is either never turned off, or the instigating stimulus of this process is constantly turning it on, inappropriately.

When this happens, there is an abnormal accumulation of collagen in the organ, in our case the bladder, and this disrupts the normal tissue architecture and function. It is normally accepted that there is an increase in Type I collagen in fibrosis that occurs in the lung (pulmonary fibrosis), the liver, and fibrotic disorders of the skin, such as scleroderma and keloids. However, recent studies have shown that in other types of fibrotic diseases, such as ischemic cardiomyopathy, there are alterations in Type III collagen. In the bladder, as in other organs, fibrosis is a chronic process. The bladder wall gradually becomes thickened and less compliant; i.e., it becomes more stiff. It is unable to cycle properly, and stores decreased volumes of urine at higher pressures.

We studied a well characterized pediatric patient population in an attempt to determine if the fibrosis of the bladder wall in these patients was the result of changes in type I collagen, type III collagen, or both.

The majority of patients had an underlying neurologic problem, i.e., they were classified as *neurogenic*: children with either a congenital abnormality (spina bifida) - a congenital abnormality of the spinal cord, in which approximately 95% of the children have abnormal enervation of the bladder, or spinal cord injury - an acquired abnormality affecting enervation of the bladder wall. A smaller population did not have any nerve related problems, but had fibrotic bladders related to bladder obstruction (posterior urethral valves, a congenital obstruction found exclusively in boys) or radiation therapy. These patients were the *non-neurogenics*. Our control patients were children who had surgery for re-implantation of their ureters.

What we found in analyzing these bladder samples was that Type III collagen protein was found in an abnormal location. Not only did there appear to be more type III collagen around the muscle bundles, but it also was found abundantly within the bundles, between individual muscle cells. We then used a biochemical assay to determine if the total amount of all collagens was different in the affected bladders. The results showed that there was an increase in the total amount of collagens (10%) in both the neurogenic and non-neurogenic bladders compared to controls.

We then carried out additional tests to determine the amounts of each type of collagen (was there more type I collagen, type III collagen or both), and found that Type III collagen was increased approximately 48% compared to the normal, control bladder tissue samples. This increase resulted in a change in the ratio of the collagens. Normal ratio in the bladder: approximately 75% type I collagen : 25% type III collagen. Neurogenic and non-neurogenic collagen ratio: approximately 67% type I : 33% type III.

What is the meaning of all of this, why is it important and how does this relate to interstitial cystitis?

Studies on fetal bladders with neurogenic or obstructive abnormalities (Ellen Shapiro, Workman and Kogan, respectively) have shown that, in utero (and therefore in the absence of any infection or inflammation) the bladder wall becomes thickened. Our data shows that this fibrosis continues during postnatal development. There is an increase in the total amount of collagen and a shift from the normal balance of specific collagen types and an increased expression of type III collagen in a location where it is not normally found.

These changes in the bladder wall proteins result in a thickening of the bladder wall along with hypertrophy of smooth muscle cells, such that it is difficult for the bladder to expand and empty properly. It also results in a bladder which has a smaller volume capacity - The bladder holds less urine, and does so at higher than normal pressures, which, over time, can lead to damage of the upper tracts (kidneys).

Another problem is that, after birth, the children are more prone to urinary tract infections and inflammation, since one of the treatments is CIC (clean intermittent catheterization). Factors which are released by inflammatory cells (cytokines, growth factors) can also cause cells of the bladder wall to increase their collagen synthesis, therefore adding to the problem. Once we understand what collagens are altered in this fibrotic process and what may be the mechanism of regulating this increased expression, then we can begin to look to interventional therapies.

Relative to Interstitial Cystitis:

A subset of IC bladders become thick walled and fibrotic and function at higher pressures. Some of the differences between the pediatric patients and IC patients is that IC, as far as I am aware, is not a congenital abnormality. However, IC is a chronic disease, characterized by inflammation, and sensory symptoms of pain, urgency and frequency. It could be possible that factors released by inflammatory cells or mast cells or inappropriate signals from nerves to the muscle cells may have an effect on the fibrotic process. Interestingly, no one has ever analyzed bladder wall tissue from IC patients to determine if there are changes in collagen expression in the thickened bladder walls. We need to do this research!

-------------------PRESENTATION ENDS---------------

<icnmgrjill> Fascinating Pam! It's so important we find research which studies the basic functions of the bladder and how a mechanism, like IC, can alter it. It reminds me of the research that we discussed a few months ago which talked about how NO2 was used as a neurotransmitter by different parts of the bladder in different ways. So, the more that we can understand about structures, the better prepared that we can be in trying to reverse or prevent that scarring, right?

<pam> That's correct. One of the first things to do is determine just what is the normal structure of the bladder. Also, developmentally it is important to know how the bladder is, basically, put together. When we know what the normal process is, then we can look at the abnormal and try to determine just what the defect is and where things went wrong.

<icnmgrjill> What types of the body actually develop fibrosis? or that stiffening??

<pam> Some of the diseases associated with fibrosis are lung fibrosis, liver fibrosis, tightening of the skin called scleroderma, sclerosis of the kidneys.

<icnmgrjill> Can patients with injuries.. like from car accidents, or with broken bones, develop fibrosis in nearby tissues?

<pam> Generally, no. However, if something does not heal properly, there can be abundant scarring which is the beginning of fibrosis. That brings me to mention that fibrosis can also be defined as abnormal wound healing. By this I mean that if there is an injury, the normal wound healing process starts. Collagens are laid down to repair the wound and then the process is shut off. However, in fibrosis, the process is either never shut off or the initiating stimulus continues, constantly turning it on, and the collagen continues to be laid down, inappropriately.

<pam> When this happens, there is thickening of the "matrix" in the walls of the organ. In the lung, the patients have a hard time breathing and exvchanging oxygen. In the kidney, it is hard to filter the blood to concentrate urine. In the bladder, whatever the stimulus is, the collagen gradually continues to accumulate, making the function more difficult.

<icnmgrjill> Is there any research that shows that IC patients have fibrosis occuring in the bladder?

<pam> Studies have stated that the bladder wall has thickened at meetings. I have seen slides from patients bladders showing what I would call fibrosis. The researchers generally say that the "connective tissue" hae appeared to thicken and basically, that is the extent of what I know has been shown.

<icnmgrjill> This is such a chicken versus the egg dilemma. Which comes first and which comes later. I think it would be fair to say that some IC patients have moderately reduced bladder capacities. I certainly did as a teenager when I had my first bladder problems. So, I guess I'm stumped here. On one hand, it makes me wonder if we have the presence of a long term inflammation which is just causing a cascade of effect of changes in the bladder (i.e. like something environmental). Or, as a few of us have conjectured, it almost makes me wonder if IC patients, for some reason, have an incomplete healing mechanism, possibly genetic in nature.

<pam> First, I think that you are right about the long term inflammation. I think that fibrosis is a gradual, gradual process. It occurs relatively slowly, probably depending upon what is the stimulus. A German reseacher presented some slides at the IC meeting in Virginia a few years ago and he said that he followed the process on slide, from biopsies, and that it was a slow gradual accumulation of "connective tissue" between the muscle of the bladder wall. The other thing about environmental in the lung, exposure to asbestos will cause the development of fibrosis if you remove the exposure, the process does not get worse.

<icnmgrjill> It makes me wish that we had some historical data that went back a few centuries that we could look at. We know that an IC like disease was first discussed in the middle 1800's and so it kind of makes me wonder if, perhaps in the onset of the industrial age, some environmental trigger began.

<pam> That a good point. It's hard to prove but I think that it is a good idea to begin to collate data about the different fibroses in different organs, look at what may be the initiating factors seen if there are any parallels.

<icnmgrjill> Pam, in your research, have you gotten into whether any of the other bladder diseases also have a reduction in bladder capacity? i.e. like radiation cystitis, or overactive bladder and, even, incontinence patients?

<pam> One of our patients was treated with radiation therapy. This patient had a thickened wall, increased pressure and increased collagen content. Another one of the patients was treated for voiding dysfunction, and this one was harder to figure out. At present, we have done nothing with incontinence patients.

<icnmgrjill> Can the prostate develop fibrosis?

<pam> Radiation definitely has an effect. Can the prostate develop fibrosis? Good question. The prostate becomes enlarged with BPH, which is benign prostatic hyperplasia. Hyperplasia is the increase in the number of cells present within the postate. We have actually tried to analyze extremely small biopsies of prostate tissue from BPH patients. However, there was just too little tissue to get a definintive answer. However, BPH can lead to what is known as outlet obstruction in men. and this could be very similar to the outlet obstruction that was seen in the patients examined in the non-neurogenic category. Older men with outlet obstruction, can develop thick walled high pressure bladders. So the mechanism may be similar in these cases.

<icnmgrjill> We know that some IC patients, including myself, have, at one point in time, also had a urethral stricture, which makes me wonder if it's possible for just a small area of the urethra to become inflammed and fibrotic?

<pam> We have examined stricture from pediatric patients. The strictures were characterized by an increase in collagen, however this collagen was type I, not the type III that we saw increased in the bladders. Could this be a predisposing condition? I really don't know, however, I intend to find out from the pediatric urologist.. if this could possibly lead to further problems in the future.

<icnmgrjill> Time is flitting by. We have time for just a few more questions. DonnaRenee brings up an interesting point. I was wondering if there has been any research done on environmental contaminants (i.e. petroleum, etc.) as to whether it can provoke reactions like fibrosis. Are you aware of any?

<pam> I am not aware of any research done on environmental contaminants. I think it is a very good idea to look into this matter

<icnmgrjill> ICN Terri wants to know if fibrosis can have a different color upon examination.. i.e. like a yellowing of the tissues?

<pam> I think it is possible. When they scope they bladder, they sometimes mention white scars. This is probably some form of scarring or fibrosis in the luminal area of the bladder. The deeper form, in the muscle, probably cannot be seen this way. However, it may be possible, don't quote me, to do ultrasound to determine the increased thickness. Whether this would show the fibrosis, I don't know, but I'll find out for you from my ped uros.

<icnmgrjill> Bev J wants to know if your research is differentiating between different regions of the bladder.. i.e. can the trigone have more or less fibrosis?

<pam> Yes, we can differentiate different regions of bladder wall relative to layers - urothelium, underlying lamina propria or what is known as mucosa and then the out muscle. The tissue sample we received were from the dome of the bladder, not from the trigone.

<icnmgrjill> Is there a way.. of slowing or stopping the fibrotic process? In IC, for example, patients who have a reduced bladder capacity are often encouraged to do bladder holding protocols to stretch out the bladder and increase capacity.

<pam> Whether exercise is right that should be tried. Maybe the collagen is being put down in the wrong orientation relative to other collagens. I don't know. However, one thing is that collagen doesn't stretch. But there is a little play to it. However, I don't think this will slow down the fibrosis. There are some interesting therapies now being developed to halt the fibrosis. Some of these are use of relaxin, to inhibit collagen over expressing, use of interferons and a new substance called halofuginone, which stops the synthesis of type I collagen, at least in the studies done.

<icnmgrjill> Are there any clinical studies being done yet?

<pam> This substance called halofuginone is now in phase II trials for treatment of scleroderma, a fibrosis on the skin.

<icnmgrjill> Terri has the pleasure of the very last question. Would repeated stretching of tissues cause fibrosis too?

<pam> That's a great question. We have done experiments on bladder cells in vitro whereby we actually stretch the cells and then look to see whether there is any change in collagen synthesis. In fact, depending on how the stretch is applied and how rapidly it is applied, we can get changes in collagen output. So yes, stretch does affect collagen output. Normal stretch is fine, but abnormal input of stretch to either smooth muscle cells or fibroblasts can alter their function.

<icnmgrjill> Pam, thank so much for being here tonight! We appreciate the presentation that you gave and the answers to our questions were certainly very thought provoking for us all. We so appreciate having the chance to see inside the workings of IC research! You were great!

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The opinion of the speaker is not necessarily the opinion of the IC-Network. Copyright © 1999, The IC Network, All rights reserved. This transcript may be reproduced for personal use only. If you do so reproduce, we ask only that you give credit to the source, the IC Network, and speakers, Pamel Howard and Jill Osborne. For additional use, please contact the ICN at (707)538-9442.