Tag Archives: macrophage

tuberculosis and edam

Robbie was really happy to be diagnosed with pulmonary tuberculosis. Before he arrived he was convinced he had lung cancer.

Robbie is in his late 50’s, he was born in Glasgow and left school at fifteen with no qualifications. His father was a shipbuilder on the Clyde. Robbie thought he would do the same, but in the seventies the shipyards closed* and there were no prospects for young men like him, so he came south.

He got various unskilled jobs, mainly in the china clay industry, got married and had two daughters. But he drank much too much and was not faithful to his wife, who left him. He says he does not entirely regret it – he had a great time.

Or as George Best famously said “I spent a lot of money on booze, birds and fast cars, the rest I just squandered”

Nationaal Archief Fotocollectie Anefo
Nationaal Archief Fotocollectie Anefo

Then, about three months ago, he developed a cough that kept him up at night. He had terrible sweats and would wake up in the morning with his bedclothes really wet. He took the sheets off his bed to hang them up to dry during the day. He knew something was seriously wrong when he started coughing up blood and was losing a lot of weight. He had always been “wiry” but now was becoming skeletal. Robbie became increasingly sure he had lung cancer, but was too frightened to get help. He still talked to his daughters every Sunday on the ‘phone. Last Sunday one of them was so worried when he told her what was happening that she went to find him and brought him in to our hospital. He had a chest Xray and within a couple of hours we told him that we thought he might have TB.

pulmonary TB usually affects the upper part of the lungs
pulmonary TB usually affects the upper part of the lungs

We put him in a side-room and got him to cough up some blood-stained sputum which we sent to the lab. Within a couple more hours we got a phone call to say that the sample was “teeming” with AFBs – acid-fast bacilli.

I have previously talked about bacterial cell walls and  the Gram stain, which is a process which involves staining a biological sampe with a blue and red stain which have different affinities for the various components of a bacterial cell wall. Gram-positive bacteria stain bluish-purple with this stain because their cell wall is mainly peptidoglycan. Gram-negative have a thin layer of peptidoglycan, but a thick covering layer of lipopolysaccharide (LPS), which, like peptidoglycan, helps protect the germ from damage. As its name suggests, LPS is made of chains of sugar molecules and lipids – with carbon chains about 16-20 carbon atoms – similar to those in triglycerides in butter and olive oil. Gram-negative bacteria look red under the microscope after the Gram staining process

Image from Centers for Disease Control and Prevention
Tuberculosis bacilli stained with auramine and counterstained with acridine orange – viewed under uv light the mycobacteria shine out –  Image from US Centers for Disease Control and Prevention –

Tuberculosis is caused by the bacterium mycobacterium tuberculosis. It has a quite different cell wall structure which makes it very difficult for our white cells to destroy.  The secret weapon is mycolic acid. This a long molecule composed mainly of hydrocarbon chains – up to 54 carbon atoms long.

Hydrocarbon chains are common in all sorts of lengths in nature, most of which are useful to us. One carbon atom, surrounded by four hydrogen atoms is methane – a gas which I rely on to cook my dinner. Three or four carbons and we get the gases propane and butane, also useful in cooking the barbecue. Octane, a liquid with eight carbon atoms gets me to work in the morning. When we get to 16-20 carbon atoms there are endless things I can get from the supermarket, mainly to eat, with this size of hydrocarbon chain. These are mainly liquids or soft solids such as olive oil, butter, meat fat, and cooking oil. With longer hydrocarbon chains we are talking about waxes. Waxes have huge numbers of use in nature and are used extensively by humans, but mainly for protection rather than as a food. We use waxes a lot for protection – shoe and furniture polish for instance.

wax is used to protect shoes, furniture and a whole lot of other things
wax is used to protect shoes, furniture and a whole lot of other things

The wax in our ears is a mixture of squalene, a hydrocarbon of 26 carbon atoms mixed with cholesterol and dead skin cells, also designed to protect us from microbial infection.  Plants produce waxes to protect the surface of their leaves, and sheep produce waxes to protect their wool (lanolin, chemically similar to earwax).  Waxes are protective because they are difficult to break down chemically. The tuberculosis bacteria use this property of wax to protect them from the nasty chemicals white blood cells use to try to destroy them. We can detect mycobacteria by using a special stain which is taken up by the waxy coating. The original stain was called Ziehl–Neelsen, or ZN, but we now use a better one called auramine. This is a fluorescent dye which sticks to the wax, even when the specimen is flooded with acid and alcohol, which will remove it from just about any other bacteria which do not have mycolic acid in their cell wall. The specimen is then viewed under ultraviolet light and the TB germs light up.

Pulmonary TB is transmitted from one human to another mainly by coughing – producing a fine spray of droplets which contain the tuberculosis bacteria. Robbie mixed with a number of alcoholics and drug addicts, who are more prone to develop this disease. When the bacteria are detected by the cells lining the lung (see asthma and pineapple below), neutrophils will have a go at engulfing them and soaking them with lysozyme and bleach. TB germs will laugh at that (no, not really, TB germs don’t have much of a sense of humour). Their waxy coat makes them impervious to hypochlorite. Then the professionals are called in –macrophages. They will also engulf the tubercle bacilli. They will then call in help from T-lymphocytes, walling off the tubercle germs in a special structure called a granuloma.

cartoon of a tuberculous granuloma - the centre is caveating - cheese-like - made from dead macrophages
cartoon of a tuberculous granuloma – the centre is caseating – cheese-like – made from dead macrophages

The TB germs can only grow very slowly, because they have to make the very long hydrocarbon chains to protect the new bacterium after cell division. Macrophages and lymphocytes use all the weapons at their disposal to try to kill them, many of which I think we don’t properly understand. We do know that nitric oxide, made by neutrophils is important. This is a gas which can penetrate the waxy coat and damage the TB germs. Another important molecule is thought to be granulysin made by cytotoxic T-cells. We don’t know how either of these molecules really do the damage and kill TB.  Usually the human wins, but in about one in ten, as in Robbie’s case, TB wins. Robbie’s smoking and previous exposure to clay dust will have caused lung damage, which makes him more susceptible. His heavy drinking and poor diet will also make it worse. Some think vitamin D, which is made in the skin from cholesterol and sunlight might be important in defence against TB. Robbie likes dim, smoky rooms and avoids sunlight most of the time. Before effective drug therapy, sanitoriums would encourage patients to sit in balconies in the sun as part of their therapy, making lots of vitamin D.

Even when the human wins, it is not always a total victory. Sometimes there is an impasse, where the TB germs cannot be killed, but only constrained in the granuloma. In this siege scenario macrophages have another trick up their sleeve – calcification. Macrophages are very closely related to cells which make bone – osteoblasts. When the granuloma cannot be sterilised the macrophages start laying down insoluble calcium salts and imprison the TB germs. Often we see white spots on the chest xrays of elderly people who were exposed to TB when they were young – this is due to the TB calcium prisons which allowed their lungs to solve the problem of what to do with indestructible organisms.

It is quite likely that Robbie will recover from his TB, as long as he takes his drug therapy regularly, even though his lungs will end up even more damaged than they were. There are some very drug-resistant strains of TB emerging in India and Africa, but at present most TB in the UK is sensitive to rifampicin, isoniazid and ethambutol. He has promised his daughter that he will stop smoking and drinking so much, and eat better food.

This week’s food link is Edam cheese.

Edam cheese is case in wax to protect it from drying out and from spoilage. I think it is too bland, but my son likes it
Edam cheese is case in wax to protect it from drying out and from spoilage. I think it is too bland, but my son likes it

This is traditionally encased in a wax coating to protect it from drying out and from microbial spoilage. I do not really understand why micro-organisms have not yet worked out how to easily break down wax and use its energy for their benefit. It has the same calorific value as fat but is clearly unappetising for all the normal germs in my kitchen which are happy to feed on shorter chain hydrocarbons.

*There is a very powerful speech by Jimmy Reid, a Glasgow shipyard worker who was elected Rector of Glasgow University.

“The greatest speech since President Lincoln‘s Gettysburg Address

– required reading:

http://www.gla.ac.uk/media/media_167194_en.pdf

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asbestos and black pudding

Some cancers are better than others. Mesothelioma is one of the worst. Alfred was admitted to our ward earlier this week. He is a 78 year old retired shipbuilder. He has mesothelioma in his lung. He spent most of his life in the Naval shipyards welding large plates of steel together to make warships. When he was young, a huge amount of asbestos was used in naval ships. It was packed in thick layers between the compartments of the vessel as it was being constructed. Alfred and his mates played snowballs with lumps of the stuff (of course, no masks were worn then).  Asbestos was used in naval construction because it is a refractory material. That means that it will not burn or melt even at high temperatures, so that it will prevent fire spreading from one part of the ship to another. It is also very cheap – a mineral that needs no further processing once it has been dug out of the ground.

Slide2
chrysotile asbestos
from Ra’ike – wikimedia commons

The reason Alfred came into hospital was because he had suddenly become more breathless and had bad chest pain when he breathed deeply. When we examined him it was clear that he had a large amount of fluid between his lung and chest cavity – a pleural effusion. We were also worried that he may have a pulmonary embolus. Patients with cancer commonly develop blood clots in their veins that travel to the lungs and block up the circulation causing shortness of breath, chest pain and even death if the clots are big enough.

His wife was with him. She knew he had cancer. She did not say it, but clearly she thought he was about to die.

We gave Alfred oxygen, an injection of low-molecular-weight heparin to prevent further blood clot formation and arranged a CT pulmonary angiogram.

Why is asbestos so dangerous?

The common form of asbestos is chrysotile. This is white asbestos. There is also brown, and the even more deadly blue asbestos. Chrysotile is a fairly simple chemical compound – magnesium silicate. Talcum powder is also magnesium silicate, but asbestos has a different crystal structure to talc. The crystals which asbestos is made of are very long, and very thin and break up into tiny, sharp needles. These needles get stuck into the lung and cannot be removed.

Ordinary dust commonly contains silicates. Sand is pure silicate. Silicon and oxygen are by far the two most common elements in the earths crust. Lots of stuff we use every day is made of silicates, including glass, ceramics (the cup you are drinking coffee out of right now), the bricks your house is made of, the cement mortar holding the bricks together, the surface of the paper you put in your printer, toothpaste etc. etc. The reason silicates are so useful is that the silicon-oxygen bond is very strong. This means when you put strong acids, alkalis or solvents in a glass vessel it does not dissolve. It also means that when it is in the wrong place in your body it can cause a problem because it cannot be broken down.

the granite worksurface, the ceramic tiles and grout, the pottery bowl, the glass and surface of the paper are made of silicates.
the granite worksurface, the ceramic tiles and grout, the pottery bowl, the glass and surface of the paper are made of silicates.

Ordinary dust, in reasonable amounts we can cope with. If it gets down into our lungs, it is trapped by the mucus layer on the surface of the tubes, or bronchi. This mucus is continually produced by goblet cells – called that because they have a goblet shape.

Mucus is a very wonderful substance. It is mainly water, but clearly not only water because it is very sticky. It is designed to be sticky to trap dust particles and bacteria. Bacteria don’t like to be trapped in mucus because they find it hard to move around – like us trying to swim in a swimming pool full of treacle.

treacle

Once the bacteria and dust particles are trapped, tiny, beating cilia on the surface of the bronchial epithelial cells move the mucus layer along. These cilia, which look like miniscule hairs, push the mucus, with its cargo of dust and germs, in one direction – upwards. Eventually, it reaches the larynx and then goes down the tunnel of death – the oesophagus – to end up in the acid of the stomach. No germs can survive this apart from helicobacter pylori – the subject of a future blog no doubt. The cilia are easily damaged. One reason smokers cough is because their damaged or absent respiratory cilia do not move mucus up the escalator, so it collects and needs to be coughed up.

Mucus is a wonderful substance made mainly of sugars. Sugars are essentially sticky substances. Lollipops are sticky, honey is very sticky. In earlier times wallpaper was stuck to walls with flour and water paste – wheat flour is composed mainly of starch –  a long polymer of the sugar, glucose.

wallpaper paste

Modern wallpaper paste is made of methylcellulose – a form of cellulose that dissolves in cold water to make something that looks quite like mucus. We use a lot of K-Y jelly in our work. It is also made from methylcellulose. I have read that many litres of K-Y jelly were used to simulate mucus in the Alien films.

The sugars in real mucus are quite special – they are in the form of long sugar chains known as mucopolysaccharides or glycosaminoglycans. I prefer the word mucopolysaccharide. It just means polysaccharide – a sugar chain – derived from mucus. They are special because unlike the common sugar glucose, many of the sugars in mucopolysaccharide chains have an amino group (NH2) and many have sulphate (SO4) groups attached. The sulphate groups make mucus even stickier.

The mucus produced by goblet cells is not just a solution of mucopolysaccharides. It is a complex structure. The goblet cell extrudes a long protein molecule called mucin. The sugar chains are attached to the mucin like bristles on a bottle brush.

Slide1
structure of mucus
the central red shaft is a protein to which many long sugary mucopolysaccharide chains are attached (blue)

So, on average, mucus has more than twice as much sugar as  protein in a large amount of water – rather like wallpaper paste.

Our bodies use mucopolysaccharides for a lot of things apart from making mucus. They are important in lubricating joints and are found in the jelly-like vitreous humor in our eyes. Some bacteria produce these sugars as a slime-coating or capsule which makes them more difficult for phagocytic cells to ingest. (See phlegm and horseradish previously).

Back to the asbestos particles. Being very thin and sharp, they manage to penetrate the mucus layer and get into the lung substance. They would not cause too much of a problem if it were not for the police in the form of macrophages. These officers of the law are very intolerant of foreign invaders and try to destroy them. (I’m not suggesting police are racist by the way). If the foreign invader is a bacterium then fair cop. If it is an organic substance like wood fibre, engulfing it and breaking it down is also a good idea. The problem with asbestos is that it is a silicate, and even though macrophages can make some pretty nasty chemicals, they cannot break down asbestos. It is like glass or ceramic. That does not stop the macrophage trying.

There is a great picture of a macrophage trying to eat an asbestos particle at the end of this imgur gallery:

http://imgur.com/gallery/nBJb6

When trying to dissolve foreign bodies, the nasty chemicals such as hypochlorite can damage DNA. This may be why asbestos exposure causes cancer, although the details are not clear at present.

There is one place in the body where foreign bodies do not cause inflammation – the anterior chamber of the eye. This is because the damage caused by cells such as macrophages would endanger our sight. The eye is known as an immunologically privileged site It means that we can put new plastic lenses in the eyes of patients with cataracts and graft a new cornea from donors without needing immunosuppressive therapy.

And now back to Alfred. He was found to have several large blood clots in his lungs. They had formed in his legs or large veins in the abdomen and travelled upwards and got stuck, reducing the flow of blood in his pulmonary arteries. We treated him with enoxaparin – a low molecular weight heparin. He got better quite quickly and went home with his wife a few days later. She looked much happier than when they arrived. We taught her how to inject her husband with heparin every day.

Heparin is a mucopolysaccharide, made from cow lungs or pig intestines. It is made from the slimy stuff on the surface of the tubes – the mucus. It is separated from the protein it is stuck to using enzymes and chemicals, and purified – the bristles are removed from the brush. We used to use this unfractionated heparin to prevent and treat blood clots.

heparin is a chain of sugars with amino groups and sulphate groups low molecular weight heparin has short chains of five sugars
heparin is a chain of sugars with amino groups and sulphate groups
low molecular weight heparin has short chains of five sugars

Now the long sugar chains are chopped up into five-sugar lengths – low molecular weight heparin. This is easier to give by an injection under the skin rather than into a vein. It is also more reliable in treating and preventing blood clots.

cartoon of fibrin breakdown - from Jfdwolff wikimedia commons
cartoon of fibrin synthesis and  breakdown – from Jfdwolff wikimedia commons

Heparin interferes with the horribly complicated cascade of events that leads to an insoluble protein – fibrin, being formed from the soluble clotting factors circulating in our bloodstream. The system is so complicated because it is vitally important. Stopping blood leaking out of holes in our blood vessels is pretty high up the list of things we have to get right if we are to survive as a species. The clotting cascade is designed to produce large amounts of fibrin clots very quickly when we need it – whether you are being savaged by a wild beast or crushed by a bus, clotting can save your life.  But blood clots in vessels without holes to repair can also be fatal. Alfred could well have died if he had not been treated with heparin.

Fibrin formation is responsible for forming clots in veins. It also strengthens the clots formed by platelets in arteries. The problem is that patients with cancer, or people who have been ill and immobile (or even after a long-haul airplane flight) can develop clots in their veins even when there is not a hole in them.

We suspected Alfred may have a blood clot problem because we found a high concentration of D-dimer in his blood. When a clot forms, say in the wall of an artery which has been damaged, it is remodelled by an enzyme called plasmin which dissolves fibrin. This shaves away the unwanted bits of clot to make them neat and fit for purpose. The fibrin turns into, among other things, D-dimer. Whenever there is a lot of fibrin clot around, the D-dimer level in the blood increases.

Now the food link. I’m particularly pleased with this one – black pudding.

black pudding
black pudding

I went to the butchers yesterday to buy some. I fried it with tomatoes, bacon and sausage for a late breakfast. Black pudding is made in the UK from pigs’ blood. It is cooked, which makes it clot, and mixed with oatmeal and herbs and stuffed into a pig’s intestine from which the inner surface or mucosa has been removed. Drug manufacturers use the mucosa to make heparin. So, black pudding is a large blood clot made in the very organ that gives us a drug for preventing blood clots!