Inflammation in Kidney Transplant Biopsies 🔬🫘

According to the Banff Classification, ten out of the 17 Banff Lesion Scores (BLS) focus on the presence and extent of inflammatory cells in different kidney compartments. These inflammatory cells, also known as leukocytes or white blood cells, are the major cellular component of the immune response, protecting the body against diseases. Therefore, they play an essential role in transplant rejection.

Leukocytes include polymorphonuclear cells (neutrophils, eosinophils, and basophils) and mononuclear cells (lymphocytes and monocytes).

For simplicity, in the MONKEY challenge, we focus on mononuclear cells (monocytes and lymphocytes). Even though pathologists do not make this distinction when performing Banff scoring, it is hypothesized that relevant information relating to disease progression may be derived from making such a subdivision, which is why we attempt to make this distinction during the challenge.

Morphology

Pathologists can differentiate different types of inflammatory cells based on certain morphological characteristics.

Monocytes are the largest cell type in the blood, with a diameter of 15-22 micrometers. They contain an ellipsoidal nucleus that is often lobulated or dented, causing a bean-shaped appearance. In comparison to lymphocytes, the nucleus usually has paler chromatin. The cytoplasm is moderate to abundant.

Lymphocytes are smaller cells with a diameter of approximately 7 micrometers. They have a rounded, dark/dense nucleus and scarce cytoplasm. 

The image below shows a case of tubulitis characterized by the presence of mononuclear cells on the basolateral aspect of the tubular epithelial cells. The mononuclear cells (arrows) are noticeable by their characteristic halo, smaller nucleus, and more condensed chromatin compared to tubular epithelial cells.

Immunohistochemistry (IHC)

However, despite typical morphological features, it can be challenging to distinguish different cell types with certainty. IHC serves as a tool to discriminate. IHC involves selectively identifying cell-specific proteins (antigens). This is being visualized by using antibodies labeled with a chromogen/reagent. We used a CD3/CD20 double staining for lymphocytes, which results in a dark brown cytoplasmic staining. The PU.1 staining for monocytes results in a nuclear magenta staining. 

The image below shows an example of interstitial inflammation in which it is more challenging to differentiate between lymphocytes and monocytes based on morphology (left image) alone. With the use of IHC (right image), it becomes clear which cells are lymphocytes (brown CD3CD20 positive cells) and which ones are monocytes (magenta PU.1 positive cells). 

Staining quality

It is important to realize that the reinforcement of IHC largely depends on the staining quality. Our multi-step restaining protocol is relatively sophisticated, and errors might occur. Also, certain tissue factors impact the restaining quality. Here are some examples of suboptimal restained sections, complicating ground-truth annotations.