Insulitis: what can be seen under the microscope in the attacked pancreas

Insulitis is the inflammation of the islets of Langerhans, the small clusters of cells in your pancreas that contain the insulin-producing beta cells. The term describes the phenomenon by which some cells of the immune system (white blood cells) gather around an islet and penetrate inside it to attack the beta cells. Insulitis is considered the hallmark of type 1 diabetes — a sign visible under the microscope that your immune system has turned against your own insulin-producing cells [1].

According to the international consensus criteria, we speak of insulitis when at least three islets each contain a minimum of 15 immune cells (marked with CD45, a protein on the surface of immune cells). The inflammatory infiltrate must also be made up predominantly of lymphocytes [1]. The number of cells involved is small, because insulitis in type 1 diabetes is a discreet inflammation, hard to observe. That is why this lesion was for a long time difficult to define and to find.

Under the microscope, an islet attacked by the immune system looks like a cluster surrounded by small, dark cells — the immune cells themselves. They crowd at the edge of the islet and sometimes penetrate inside it. Doctors distinguish two situations. In a mild attack (peri-insulitis), the immune cells stay mainly at the periphery of the islet. In an advanced attack, they spread among the functional cells inside. Most immune cells remain at the edge of the islet and relatively few directly touch the beta cells. The attacked islet stains progressively more weakly for insulin, a sign that it is losing its beta cells [2].

A characteristic feature is the islet emptied of insulin-producing cells. It nonetheless keeps its other types of cells, such as the alpha cells (which produce glucagon) and the delta cells (which produce somatostatin). These islets are smaller and no longer stain for insulin. The immune attack withdraws after the beta cells have been completely destroyed, so the emptied islets look quiet, without inflammation. In a pancreas with type 1 diabetes there is a mosaic: some islets look normal and full of insulin, others are actively inflamed, and many are emptied of beta cells, as the figure below shows [2].

The infiltrate is dominated by T lymphocytes, the most numerous being the cytotoxic T lymphocytes (CD8+), which directly attack the beta cells. Also taking part, in smaller numbers, are macrophages, helper T lymphocytes, B lymphocytes, plasma cells, rare regulatory T lymphocytes and occasionally natural killer cells [3]. B lymphocytes are rare in early insulitis and become more numerous as the destruction of the beta cells progresses. They are more abundant in children diagnosed under 7 years of age.

A surprising aspect is that human insulitis is an inflammation of low intensity. The actual number of infiltrating cells is modest, much smaller than the dramatic inflammation observed in laboratory animals or in other autoimmune diseases [3]. This discreetness is one of the reasons why insulitis was for a long time hard to recognize and was considered an elusive lesion. The attack is so weak that it can pass unnoticed on an ordinary examination.

The insulitis process is not spread uniformly throughout the pancreas, but appears in a "dalmatian" pattern, in the form of small spots. The pancreas is anatomically organized into lobules, and the disease tends to affect a whole lobule at once. A lobule full of islets emptied of beta cells can sit right next to a lobule whose islets look almost normal, full of insulin. At clinical onset (stage 3), the loss of beta cells is severe in most lobules, but neighboring lobules with an almost normal number of beta cells can persist [4].

It is not fully clarified why this happens. The main hypothesis is that the autoimmune process advances lobule by lobule, possibly because of the way the blood vessels, nerves and ducts are organized inside the pancreas [4]. This patchy distribution has an important practical consequence. A small sample (for example a biopsy) from a single area could hit a lobule untouched by inflammation and completely miss the diagnosis of insulitis.

The basic principle is that the immune system attacks the islets that still contain beta cells. Studies on the pancreas show that insulitis affects predominantly the islets that contain insulin — approximately one third of them. The islets emptied of beta cells are almost completely spared [5]. Once an islet has lost its beta cells, the immune cells lose their target and move away, leaving that islet untouched from then on.

An islet without beta cells becomes practically invisible to the immune attack and can persist that way indefinitely. In addition, there are also islets that keep their beta cells without having yet been found by the immune system. These mechanisms explain why the islets look so different within a single pancreas. The inflammation of the immune attack moves unevenly, avoiding the islets emptied of beta cells [5].

Most of the information about human pancreatic tissue comes from fragments taken from deceased organ donors, because the pancreas of a living person is very hard to examine. There are official international networks that collect pancreases from deceased donors. The donors either had type 1 diabetes or only had autoantibodies, without having yet developed the disease (stages 1 and 2). These networks distribute the tissue free of charge to researchers all over the world [6]. A historical collection also keeps tissue obtained especially from children who died close to the moment of diagnosis.

These tissue collections have shown that insulitis is real, discreet and distributed in the form of many uneven patches throughout the pancreas. Researchers could see directly that the loss of beta cells is lobular, with emptied islets coexisting with islets full of insulin. The attack targets mainly the islets that still contain insulin [6]. It should be noted, however, that the samples studied internationally come from a relatively small number of donors.

The landmark study is a research program from Norway (DiViD). The team took small fragments from the tail of the pancreas in a few young adults, just a few weeks after the diagnosis of type 1 diabetes. There was a suspicion that tissue from deceased donors, taken a few hours after death, might not faithfully reflect the living organ. These biopsies therefore offered a unique image of the living pancreas, right at the onset of the disease [7].

The study confirmed that the criteria for insulitis were met in all patients. The infiltrating cells were mainly T lymphocytes located around the islets (peri-insulitis). An important reserve of islets full of insulin was still present, some completely normal and capable of secreting insulin [7]. The program also showed the risks, however: it was stopped because of serious complications (bleeding and leakage of pancreatic juice). That is why such biopsies cannot become a routine procedure.

No. The age at which the disease appears strongly influences the way insulitis looks. In general, two patterns are recognized. In children diagnosed before about 7 years of age, insulitis is more aggressive and affects a larger proportion of islets. It is rich in B lymphocytes and is associated with a rapid, almost complete loss of the beta cells. In those diagnosed at 13 years or later, insulitis is milder, contains few B lymphocytes and leaves many more islets full of insulin intact [8]. Those diagnosed between 7 and 12 years can have the features of either of the two patterns.

There is an important practical consequence. Patients with onset at older ages keep at diagnosis a significant part of the insulin-producing islets. This suggests that in them the functional deficit matters more than the absolute loss of beta cells [8]. This difference has implications for treatment, because therapies that target B lymphocytes could act differently in the two groups. Observations from the pediatric population also show that a younger age at onset is associated with a more severe form of the disease and with more pronounced pancreatic inflammation [9].

Yes. Insulitis is most intense and most frequent close to diagnosis, especially in young people. Today it is clear, however, that the autoimmune attack is chronic and can continue for years, at reduced intensity. In tissue from donors, insulitis and surviving islets full of insulin have been found many years after diagnosis. Beta cells found together with insulitis a few years after onset show that islet autoimmunity persists well into the period after diagnosis [5].

This persistence changes the way we understand type 1 diabetes: it is not a single destructive event that ends at diagnosis, but a continuous and dynamic process. As long as islets containing insulin remain, they remain possible targets, so the immune attack continues, admittedly at very low intensity [4]. This chronic character is, however, encouraging. It means that there may be a prolonged window in which the surviving beta cells could be protected or saved by treatment.

Remarkably, yes. The Joslin Medalist Study looked at people who have been living with insulin-dependent diabetes for 50 years or more. A large proportion of them still produce a detectable amount of C-peptide, the molecule released together with insulin. This is proof that the body still makes a very small amount of its own insulin. Some of these people even respond to a meal with a rise in C-peptide. This overturned the conviction that the beta cells are destroyed completely and uniformly [10].

Examination of the pancreas in these people confirmed the observation at the tissue level too: all the pancreases examined kept insulin-producing beta cells, isolated or in small groups [10] [11]. Some cells showed both signs of cell death and signs of division. This suggests a continuous balance between the loss and the renewal of beta cells, even after decades of disease. The implication is profound. If even a few beta cells survive and renew themselves over tens of years, strategies to protect or multiply them could be useful even in those with long-standing diabetes.

There are three solid reasons. Firstly, the pancreas is a dangerous organ to biopsy, because it is full of digestive enzymes. An incision risks leakage of pancreatic juice, with inflammation of the pancreas and significant bleeding. Secondly, the patchy and lobular distribution of insulitis means that a small sample can easily hit an untouched area and may then give a false negative result. A single biopsy is therefore inconclusive.

Thirdly, the biopsy is quite simply useless for diagnosis, because type 1 diabetes is diagnosed clinically and confirmed through blood tests, such as the autoantibodies and the measurement of C-peptide. Current guidelines build the diagnosis and staging on the clinical signs, on blood glucose and HbA1c, on autoantibodies and never on examination of the pancreatic tissue [12]. Subjecting a patient to a risky surgical procedure to confirm something that a blood test has already answered would be unjustified, so the biopsy remains reserved for research.