Karl Landsteiner and the Discovery of Blood Groups

Karl Landsteiner and the Discovery of Blood Groups Core Module Immunology Doctoral Training Group GK1660 Erlangen  2011 Lena Krzyzak

Outline • Life of Karl Landsteiner • History of the Discovery of Blood Groups • Landsteiner and his Discovery of Blood Groups • Discovery of Rhesus Factor • Haemolytic Disease of the New-born • Blood Group System Today

Life of Karl Landsteiner

Karl Landsteiner (1868-1943) • born June 14th, 1868 in Baden (near Vienna) • 1885 Study of medicine at the University of Vienna (Austria), graduated 1891 • Studies in organic chemistry at the • University Zürich – Arthur Hantzsch • University Würzburg – Emil Fischer • University of Munich – Eugen Bamberger • 1891 published 1st paper on the influence of diet on the composition of blood ash • 1896 Vienna General Hospital - Assistant under Max von Gruber, Institute for Microbiology and Hygiene • 1897 Vienna Institute of Pathology - Performing autopsies 3639 dissection Speiser, 1961 (Hollinek)

Karl Landsteiner (1868-1943) • 1900 Description of the agglutination of human blood • 1901“Agglutination phenomena of normal human blood” • 1908 Department of Pathology – Wilhelmina Hospital Vienna (Head) • 1911 Professor of Pathological Anatomy in the University of Vienna • 1919 Hospital The Hague, Netherlands • 1922 The Rockefeller Institute, New York • 1929 Landsteiner became US citizen • 1930 Nobel price for medicine • 1939 Emeritus Professor at the Rockefeller Institute • Died in 1943 at the age of 75 in New York, USA Speiser, 1961 (Hollinek)

Work of Karl Landsteiner Pathogenesis of paroxysmal haemoglobinuria Human blood groups ABO Amongst the first to purify antibodies Introduction of Haptens Introduction of dark-field microscopy to visualize the spirochetes of syphilis Polio research: Polio belongs to a group of filterable microorganisms Research work on Syphilis  Detection of Treponemapallidum  360 Publications Most of the time, Landsteiner was quite poor

Nobel price for Medicine 1930 "for his discovery of human blood groups" ‘Beginning in 1923, fourteen different nominators had put him up for the award for three different discoveries - his polio research, his immune system work, and his discovery of the blood groups. After 29 years, the committee finally granted him the award for his blood group work.’ scienceheroes.com

History of the Discovery of Blood Groups

Adolf Creite (1869) • born 1847 in Helmstedt, Lower Saxony (died 1921) • Medical student at the University of Göttingen • Physiological Institute, Prof. Georg Meissner • 1869 ‘Zeitschrift für Rationelle Medizin’ • ‘Versuche über die Wirkung des Serumeiweisses nach Injection in das Blut’ • Serum proteins could „dissolving“ (lysis) and „cluster“(agglutination) • Inspired by Stokvis (1867) and Bernard (1856) Hughes-Jones, 2002 (Brit. Jour. Haem.)

Creite‘s Experiments • Sera from • calf • pig • dog • sheep • cat • chicken • duck • goat • calf • pig • dog No Effect Injection of 8 ml serum • sheep • cat • chicken • duck • goat • “blood-stained urine“ • general malaise • death • Blood-stained urine: similar observations in literature (injection of whole blood rather than serum)  Panum (1863) •  Sheel (1803) – Ippolito Magnani as well as Denis and Gaspard de Gurye Creite‘s conclusion: Constituents of serum had certain ‘chemical properties’(chemische Eigenschaften) which affected foreign red cells directly (‘dissolve red cells’)  Active ingredient could be serum protein Hughes-Jones, 2002 (Brit. Jour. Haem.)

Creite: 1st Description of Agglutination In vitro Experiments • Sera from • dog • sheep • cat • chicken • duck • goat Add to a drop of fresh blood ‘cells suddenly flow together forming different-shaped drop-like clusters’ Microscopy Possibility that some blood cells has dissolved • rabbit No Effect Creite described his observation but did not reach the conclusion that the formation of ‚drop-like clusters‘ could be a true agglutination reaction Hughes-Jones, 2002 (Brit. Jour. Haem.)

Leonard Landois (1875) • born 1837 in Münster • Medical student at the University of Greifswald • Professor and Director of the Institute of Physiology at Greifswald • Study of blood transfusions and the phenomena of agglutination • 1875 published: ‘Die Transfusion des Blutes’ • did not cite Creite • died 1902 in Greifswald Lehrbuch der Physiologie des Menschen (1880) Hughes-Jones, 2002 (Brit. Jour. Haem.)

Landois‘ Experiments Question: Is lysis of red cells are those of the recipient or the donor? Incubation at 37°C Add fresh blood Observation of the initiation of red cell lysis Sera from eight different animals Transparent Non-transparent Incubation at room temperature Transparent, cells are no longer visible Add fresh blood Incubation • Nearly all foreign red cells lyse within minutes • cells in close proximity to each other form clumps Sera from dogs Transparent Non-transparent e.g. Sera from dogs Hughes-Jones, 2002 (Brit. Jour. Haem.)

Landois‘ Experiments Question: Is lysis of red cells are those of the recipient or the donor? Incubation at 37°C Add fresh blood Observation of the initiation of red cell lysis Sera from eight different animals Transparent Non-transparent Incubation at room temperature Transparent, cells are no longer visible • High concentration of red cells  clumping of red cells occurred rather than lysis •  substance in the serum which acted on red cells and made the membrane soft and sticky, when cells touched each other they aggregate Landois was stuck with the great variation in the activity shown by different sera on different cells, but considered that lysis and aggregation resulted from some type of interaction between the serum and the red cells Hughes-Jones, 2002 (Brit. Jour. Haem.)

Landsteiner and his Discovery of Blood Groups

The Way to Landsteiner 1890 Discovery of the antibodies specific for tetanus by Emil von Behring und Kitasato 37 Papers on the subject of red cell agglutinins and lysins published 1898-1901 1900 Shattock and Grünbaum  appearance of serum agglutinins were a manifestation of certain diseases (esp. infections)  Landsteiner entered the field 1900 Landsteiner: ‘Zur Kenntnis der antifermentativen, lytischen und agglutinierenden Wirkungen des Blutserums und der Lymphe’ 1900 Ehrlich und Morgenroth  Agglutination of human blood with human serum is called ‘Isoagglutination’(Berliner klinische Wochenschrift)

Rouleaux Rouleaux are stacks of red blood cells, which form because of the unique discoid shape • Occur, when plasma protein concentration is high •  erythrocyte sedimentation rate is also increased • Non-specific indicator of the presence of diseases wikipedia.org

1900  First suggestion of the existence of serum agglutinins and red cell antigens (footnote): Landsteiner, 1900 (Wiener Klin. Wochenschrift)

1901 Landsteiner‘s 17th publication Classification of three different groups according to its agglutination properties Landsteiner, 1901 (Wiener Klin. Wochenschrift)

Crosstesting sera and red cells from six healty scientists including himself Serum of Dr. Störck and Landsteiner reacted with no other red cells None of the sera reacted with their own red cells  First description of self-tolerance Landsteiner, 1901 (Wiener Klin. Wochenschrift)

Crosstesting sera and red cells from six healty scientists including himself Serum of Dr. Pletsching reacted with Dr. Sturli cells Landsteiner suggested at least two classes of antibodies: anti-A and anti-B Dr. Pletsching  A antigen on red cells & anti-B antibodies in his serum Dr. Sturli  B antigen on red cells & anti-A antibodies in his serum Cells of Dr. Störck and Landsteiner contained neither A antigen nor B antigen, but anti-A and anti-B antibodies in their sera  3rd type: C (today: 0) Landsteiner, 1901 (Wiener Klin. Wochenschrift)

Mechanism? • Eisenberg: Resorption of red cell particles? • Landsteiner rejected this idea  he was not able to produce autoagglutinins after injection of their own red blood cells in animals ? • 2) Agglutinins forming as a result of physiological decay of organ tissue: • phenomenon of autoimmunization induced by red cell particles • linked to diseases from which people had recovered  Exact triggers responsible for the production of anti-A and anti-B in humans have yet to be identified (Historical Review, 2002)  Today: Antibodies induced due to natural exposure to similar antigen determinants on microorganisms present in the normal flora and the gut

1902: Discovery of 4th group (AB) (von Decastello and Sturli) Alfred von Decastello-Rechtwehr (1872–1960) and Adriano Sturli (1873–1964), two kollegues of Landsteiner, discovered one year later the 4th group and called it AB. von Decastelo, A. & Sturli, A. (1902) Über die Isoagglutinine im Serum gesunder und kranker Menschen. Münchner Medizinische Wochenschrift, 49, 1090-1095. 1910-1911 von Dungern and Hirszfeld Discovery of mendelian inheritance of A and B von Dungern, E. & Hirszfeld, L. (1911) Über gruppenspezifische Strukturen des Blutes III. Zeitschrift für Immunologische Forschung, 8, 526-562. 1910 von Dungern and Hirszfeld Nomenclature for the AB0-System  1928 International accepted

History of the Discovery of Blood Types 1869 1875 1901 1902 1911 Adolf Creite Leonard Landois Karl Landsteiner von Decastello and Sturli von Dungern and Hirszfeld 1st description of agglutination Research on agglutination Discovery of the blood groups A, B and C (0) Discovery of mendelian inheritance of A and B Discovery of the blood group AB Kuby 5th edition

The Rhesus factor

Rhesus factor – Discovery 1921 Unger: Report of intragroup transfusion reactions  Additional tests should be carried out for saver blood transfusions • 1939 Levine: Report of an unusual case of intragroup agglutination (Levine and Stetson, 1939) • Woman (blood type 0) with a stillbirth needed a blood transfusion from her husband (0)  within 10 min she developed severe symptoms • Cross-matched: her serum agglutinated with her husband‘s cell • 104 group 0 samples were tested  only 21 were compatible • Levine suggested: Isosensitization caused by ‘products‘ from the fetus 1940 Landsteiner and Wiener: Discovery of Rh factor ‘An agglutinable factor of human blood recognizable by immune sera for rhesus blood‘ Human alloantibody was renamed ‘anti-D‘ Landsteiner & Wiener, 1940 (Proceedings of the Society for Experimental Biology and Medicine)

1941 Immunization Blood from rhesus monkeys Obtaining serum Agglutinable factor different from A and B was detected • Clinical importance: • Patients with haemolytic reactions after receiving repeated blood transfusion of the proper blood group • Serum of these patients contained anti-Rh antibodies (but cell factor was lacking) • Appearance of immune isoantibodies in pregnancies Rhesus factor Goal: Develop a practical method of testing for the presence of Rh factor and investigate heredity

Test System for the Presence of Rh Injection of cells i.p. After 7 days Washed red blood cells from rhesus monkeys Obtaining serum Using sera for practical diagnosis Day 1 and 5 Results Sera + washed blood suspension In narrow tube wait 30 min Sedimentation Inspection of bottom of tube  Typing of patients and prospective blood donors  Examination of families showed Rh factor is inherited Mendelian dominant

Haemolytic Disease of the New-born within 24-48h after 1st delivery Kuby 5th edition

Haemolytic Disease of the New-born • Caused by destruction of red blood cells due to alloantibodies (IgG) against Rh antigens • Maternal IgG antibody passes through the placenta binding the foetal Rh-positive red blood cells and destroying them • leading to anaemia  foetal brain damage  death (erythoblastosisfetalis) • different forms: from mild to severe anaemia • Development of haemolytic disease of the new-born can be detected by testing maternal serum during the pregnancy for antibodies of the Rh antigen •  Rise in antibody titer  Coombs test Isolated fetal red blood cells + Coombs reagent (goat anti-human IgG)  Maternal IgG bound  Agglutination Treatment during pregancy: intrauterine blood transfusions, plasmapheresis of the mother‘s blood Kuby 5th edition Haemolytic disease (65%) due to AB0 incompatibility  minor consequences (elevation of bilirubin  exposureofinfanttolowlevelsof UV light)

Human blood groups – Today (2010) 308 blood group determinants 30 blood group systems  last one: RHAG (≈2008) • MNS system • 46 antigens • describedby Landsteiner and Levine (1927) • Kell-Cellanosystem • Kell protein • Alloimmuneor autoimmune diseases • McLeodsyndrome Goeff, 2001 (Wiener Klin. Wochenschrift) Nomenclature and updated tables regarding new blood groups and their antigens  International Society for Blood Transfusion

Distribution World Wide wikipedia.org

Blood Transfusions - History Report on a dog-dog blood transfusion from physiologist R. Lower  most experiments were successful 1666 1667 • 1st successful blood transfusion to a human from a lamb by two french physicians Denis and Emmerez • 4th transfusion unsuccessfull (patient got 2nd transfusion): haemolytic reaction due to immunization of the patient by 1st transfusion • Transfusion were prohibited 1907 1st successful transfusion achieved by Dr. Reuben Ottenberg (Mt. Sinai Hospital New York Today Extensive blood donor screening  Reduced risk for HIV, HBV and HCV infection as well as for AB0 incompatibility  Packed red blood cells (‚Erythrozytenkonzentrate‘) Schwarz, 2003 (Brit. Jour. Haem.)

Importance of Landsteiner’s Discovery Landsteiner (1901) Superman of Science Makes Landmark Discovery - Over 1 Billion Lives Saved So Far! Almost 4 million American receive a blood transfusion each year Worldwide, there are about 890 million units of blood donated each year. scienceheroes.com

“Wherever a blood transfusion is performed in the world today, wherever a worried mother’ s threatened child is saved, Karl Landsteiner is virtually present” Herrmann Chiari, 1961

Thank you for your attention! scienceheroes.com

Karl Landsteiner and the Discovery of Blood Groups