Monday, March 14, 2011

Perils of Poly(agglutination)

OK, here's one for all you immunohematology geeks out there (I say that with all admiration and respect, by the way)!

Not long ago, the immunohematology reference lab at the blood center where I work received a sample for a lectin workup from a patient suspected to have polyagglutination. The patient was a child, and had recently been diagnosed with sepsis secondary to Streptococcus pneumoniae. This situation led me to think about polyagglutination and summarize it for you.

Polyagglutination is poorly understood by most students and even more poorly understood by most blood bank physicians. It is a situation in which red blood cells are agglutinated in the presence of virtually all human serum as a consequence of a change from normal that occurs on the surface of the red cell. This "change" is the exposure of antigens that normally remain hidden from view, either as a consequence of an infection or as an inherited disorder.

Multiple forms of polyagglutination (also called "activation") have been described, and they share one important feature: The not-normally-exposed exposed antigens (properly called "cryptantigens") are targeted by IgM antibodies present in the circulation of the vast majority of human sera. Those antibodies cause the diffuse agglutination of activated red cells, regardless of the ABO type of the red cells or ABO or other antibodies in the serum (thus, the designation "polyagglutination").

In general, polyagglutination is seen far less often in modern blood banks than in the past. This is largely due to the fact that the majority of cases of polyagglutination were suspected when a patient or donor had an ABO discrepancy on routine blood typing tests. Such findings were due to the reaction of the polyagglutinable RBCs against the antibodies (polyagglutinins) in most human serum (remember, pooled human serum was used in the past for ABO typing). Since monoclonal reagents are used for these tests today, cases of polyagglutination are not so obvious, and they usually are  not noticed until human serum and red cells are mixed together in performance of an antibody screen, antibody panel, or crossmatch, or a suspicious clinician alerts the blood bank due to clinical symptoms.

It is easy to get overwhelmed in the sea of different types of polyagglutination, but I recommend that you just think of them in two categories: Acquired (which is most common) and Inherited.

Acquired PolyagglutinationInherited Polyagglutination
T Activation
Tn Polyagglutination
Tk, Th, Tx Polyagglutination
Acquired B
Sd(a++); Cad

Acquired Polyagglutination
The most common and best example of this type of polyagglutination is known as "T activation." Also known as the "Hubener-Thomsen-Friedenreich phenomenon" (in honor of the three researchers who discovered and figured it out), this form of polyagglutination is seen most in children, primarily in those with bacterial infections. T activation occurs most often in association with infections with Streptococcus pneumoniae, Clostridium perfringens, or Vibrio cholerae, though the same findings can be present in influenza virus infections. Enzymes from the bacteria cleave sialic acid residues (N-acetylneuraminic acid or "NeuNAc") from glycophorins A and B, exposing the underlying T antigen to full view (while also decreasing expression of MNS blood group antigens carried on the same glycophorin chains). Most of these patients have no symptoms, but occasionally, someone with T activation may present with hemolysis, even without being transfused (it is theorized that RBCs with T activation may be cleared much more rapidly from the circulation). More significantly, patients with T activation may suffer significant hemolysis when they are transfused with human plasma. Patients with T activation may require the use of washed cellular blood products to avoid hemolysis, but such use is transient, as this antigen change only lasts until the infection is cleared. T activation has a characteristic lectin reaction pattern (outlined below).

Tn polyagglutination is a classic polyagglutination that is acquired, but NOT acquired from an infection. It is caused by a mutation on a gene on the X chromosome that leads to poor synthesis of sialic acid residues on glycophorins A and B (with a corresponding MNS antigen decrease). As a result of the glycophorin A and B/MNS connection, it is easily confused with T activation. However, the two entities differ in several important ways. First, Tn polyagglutination is associated with much more serious clinical problems than T activation, including the common associated findings of hemolytic anemia, thrombocytopenia, and leukopenia. Second, Tn polyagglutination is lifelong (since it is caused by a mutation), as opposed to the transient nature of T activation. Third, the red cells in Tn polyagglutination are not uniformly agglutinated, since some of the RBCs came from the mutated clone and others did not, leading to a characteristic "mixed field" agglutination pattern when mixed with human serum. Finally, both T and Tn cryptantigens may be over-expressed on malignant cells, but high Tn density on those cells has been associated with higher metastatic potential.

The other acquired polyagglutination entities outlined have unique features, but are all associated with characteristic infections and resultant cryptantigen exposure.

Inherited Polyagglutination
HEMPAS is an inherited hemolytic anemia in which the red cells undergo polyagglutination, associated with an antibody that can cause hemolysis of affected red cells at 37C. Sd(a++) cells (also fabulously named "super-Sid" or "Cad") are seen when individuals have an abnormally strong form of the Sd^a antigen, leading to polyagglutination. Individuals harboring these cells are not reported to show substantial resultant adverse effects.

Lectin Reactions
Lectins are at the core of how blood banks work up suspected cases of polyagglutination. It is important to note, however, that lectin reaction patterns are not to be used as the only arbiter of a specific type of polyagglutination. Clinical and laboratory data are also a large part of the workup. With that being said, here are some classic reactions with various lectins, patterned after Daniels, Human Blood Groups, 2nd ed., page 525, and Reid, The Blood Group Antigen Facts Book, 2nd ed., page 545.

Lectins T ThTnHEMPASCad
Arachis hypogea++000
Glycine max (soja)+0+00
Salvia sclarea00+00
Salvia horminum00+0+
Dolichos biflorus00+0+

Many thanks to Monica LaSarre and Colleen Chiappa, two of the geniuses at the Bonfils Blood Center Immunohematology Reference Laboratory, who helped fact check this post. Anything smart written here should reflect their brilliance, while any errors are surely mine.

Updated June 2014 to correct categorization error with Tn polyagglutination.