90 CHAPTER 2 Red cell disorders Diagnosis Demonstrate enzyme defi ciency. In suspected RBC enzymopathy, assay G6PD and PK f i rst, then look for unstable Hb. Diagnosis is diffi cult during haemolytic episode since r eticulocytes have iilevels of enzyme and may get erroneously normal result; wait until stea dy state (76 weeks after episode of haemolysis). Family studies are helpful (see Fig 2.12). Management \200 Avoid oxidant drugs \200\224see BNF1. \200 Transfuse in severe haemolys is or symptomatic anemia. \200 IV fl uids to maintain good urine output. \200 exchan ge transfusion in infants. \200 Splenectomy may be of value in severe recurrent haemolysi s. \200 Folic acid supplements (?proven value). \200 Avoid Fe unless defi nite Fe defi ciency. GLUCOSE ATP Glucose 6-phosphate dehydrogenase ADP Glucose 6-phosphogluconate Glucose 6-phosphate NADPH NADP Hexose monophosphate shunt Fructose 6-phosphate ATP ADP GSSG G-SH Fructose 1,6-diphosphate Ribulose 5-phosphate Glyceraldehyde 3-phosphate NAD GLYCOLYSIS NADH 1,3-diphosphoglycerate Mutase ADP 2,3-diphosphoglycerate ATP 3-phosphoglycerate Phosphatase Luebering-Rappaport Shunt 2-phosphoglycerate Phosphoenolpyruvate ADP Pyruvate kinase ATP Pyruvate Fig. 2.12 Glycolytic pathway showing key enzymes in red. Reference 1. British National Formulary. M www.bnf.org Further reading Mason, P.J. et al. (2007). G6PD defi ciency: the genotype-phenotype associa tion. Blood Rev 21, 267 \200\22383. 91 PYRUVATE KINASE DEFICIENCY Pyruvate kinase de \201 ciency Congenital non-spherocytic haemolytic anemia, caused by def i ciency of PK enzyme (involved in glycolytic pathway), leading to unstable enzyme with red uction in ATP generation in RBCs. O2 curve is shifted to the right due to i 2,3-DPG product ion. Epidemiology Autosomal recessive. Aff ected persons are homozygous or double het- erozygote s. Clinical features Highly variable, with chronic haemolytic syndrome. May be apparent in neo nate (if severe) or may present in later life. Diagnosis \200 Variable anemia. \200 Reticulocytes ii. \200 DAT \210\222ve. \200 L DH i. \200 Serum haptoglobin d. \200 Defi nitive diagnosis requires assay of PK level. Complications Aplastic crisis may be seen in viral infection (e.g. parvovirus B19). Treatment Dependent on severity. General supportive measures include daily folic acid (5mg/ d). Transfusion may be required. Splenectomy may be of value if high transfusion requiremen ts. In aplastic crisis (e.g. viral infec- tion) support measures should be used. Further reading Prchal, J.T. et al. (2005). Red cell enzymes. Hematology Am Soc Hematol Edu c Program, 19 \200\22323. Zanella, A. and Bianchi, P. (2000). Red cell pyruvate kinase defi ciency: from genetics to clinical manifestations. Baillieres Best Pract Res Clin Haematol, 13, 57 \200\22381. 92 CHAPTER 2 Red cell disorders Other red cell enzymopathies Glycolytic pathway \200 Hexokinase defi ciency. \200 Glucose phosphate isomerase defi c iency. \200 Phosphofructokinase (PFK) defi ciency. \200 Aldolase defi ciency. \200 Tr iosephosphate isomerase (TPI) defi ciency. \200 Phosphoglycerate kinase (PGK) defi ciency . Epidemiology Incidence <1 in 106. Inheritance is autosomal recessive (most double heterozyg ote) except for phosphoglycerate kinase defi ciency (X-linked recessive). Clinical features Similar to PK defi ciency although most are more severely aff ected for t he degree of anemia (glycolytic block results in d 2,3-DPG and left shift of O2 dissociatio n curve). PFK defi ciency is associated with myo- pathy. TPI and PGK defi ciencies are asso ciated with progressive neuro- logical deterioration. Diagnosis \200 E See Pyruvate kinase defi ciency, p.91. \200 Non-specifi c morphology w ith anisocytosis, macrocytosis, and polychromasia. Defi nitive diagnosis requires assay of defi cient enzyme (lreference lab). Complications \200 E see Pyruvate kinase defi ciency, p.91. Treatment Folic acid (5mg/d). Transfusion may be required (beware Fe overload if high trans fusion requirement). Role of splenectomy controversial. Natural history Similar to PK except TPI and PGK \200\223TPI present in childhood and cause progressive paraparesis, most die <5 years old due to cardiac arrhyth- mias. PGK can cause exertional rhabdomyolysis and consequential renal failure. Those aff ected show progressiv e neurological deterioration. Nucleotide metabolism \200\224pyrimidine 5 \200 nucleotidase de \201 ciency Epidemiology Autosomal recessive. Note: lead poisoning causes acquired pyr imidine 5 \200 nucleotidase defi ciency. 93 OTHER RED CELL ENZYMOPATHIES Clinical features \200 Moderate anemia (Hb 710g/dL). \200 i reticulocytes. \200 i bil irubin. \200 Splenomegaly. Diagnosis \200 RBCs show prominent basophilic stippling. \200 Pyrimidine 5 \200 nucleo tidase assay. Treatment Symptomatic, splenectomy is of limited value. 94 CHAPTER 2 Red cell disorders Drug-induced haemolytic anemia Large number of drugs shown to cause haemolysis of RBCs. Mec hanisms variable. May be immune or non-immune. \200 Some drugs interfere with lipid compo nent of RBC membrane. \200 Oxidation and denaturation of Hb: seen with, e.g. sulfonamides , especially in G6PD-defi cient subjects, but may occur in normal subjects if drugs given i n large doses, e.g: \200 Dapsone. \200 Sulfasalazine. Hapten mechanism describes the interaction between certain drugs and the RBC membrane components generating antigens that stimulate antibody production. DAT +ve. \200 Penicillins. \200 Cephalosporins. \200 Tetracyclines. \200 Tolbutamide. Autoantibody mediated haemolysis is associated with warm antibody mediated AIHA. DAT +ve. \200 Cephalosporins. \200 Mefenamic acid. \200 Methyldopa. \200 Procainamide. \200 Ibuprofen. \200 Diclofenac. \200 Interferon alfa. Innocent bystander mechanism occurs when drugs form immune complexes with antibody (IgM commonest) which then attach to RBC membrane. Complement fi xa tion and RBC destruction occurs. \200 Quinine. \200 Quinidine. \200 Rifampicin. \200 Antihistamines. \200 Chlorpromazine. \200 Melphalan. \200 Tetracycline. \200 Prob enecid. \200 Cefotaxime. Laboratory features As for autoimmune haemolytic anemia, d Hb, i reticulocytes, etc. Diff e rential diagnosis \200 Warm/cold autoimmune haemolytic anemia. \200 Congenital haemolyt ic disorders, e.g. HS, G6PD defi ciency, etc. Treatment \200 Discontinue off ending drug. \200 Choose alternative if necessary. \200 If DAT +ve with methyldopa no need to stop unless haemolysis. \200 Corticosteroids generally unnecessary and of doubtful value. \200 Transfuse in severe or symptomatic cases only. \200 Outlook good with complete reco very usual. 95 METHAEMOGLOBINAEMIA Methaemoglobinaemia The normal O2 dissociation curve requires Fe to be in the ferrous form (i.e. reduced, Fe2+). Hb containing the ferric (oxidized, Fe3+) form is termed methaemoglob in (MetHb). MetHb is unable to bind O2 leading to poor tissue oxygenation. In health metHb should be \211 3% total Hb. May be congenital or acquired. (See Table 2.12.) Table 2.12 Methaemoglobinaemia Methaemoglobinaemia Congenital HbM or globin mutation in vicinity of Fe. Fe becomes stabilized in Fe3+ form. Heterozygote has 25% HbM MetHb reductase def. Due to defi ciency of NADH-cytochrome b5 reductase. Autosomal recessive inheritance; sympto ms mainly in homozygote Cyanosis from infancy. PaO2 is normal. General health is good Clinical features Occurs when RBCs are exposed to oxidizing agents, producing HbM. Implicated agents include: phenacetin, local anesthetics (e.g. lignocaine), inorganic nitrates (NO2). Patients may ex perience severe tissue hypoxia. HbM binds O2 tightly and fails to release to tissues Acquired 22 HbM = 50% requires urgent medical attention. Diagnosis May be history of exposure to oxidant drugs or chemicals. Spectro- photometry or hemoglobin electrophoresis will demonstrate HbM. Assays for MetHb reductase are available. Treatment In patients with congenital symptomatic HbM give ascorbate or meth- ylthioninium chloride (methylene blue). In acquired disorder remove oxidant, if present, and administer methylthioninium chloride (meth- ylene blue). 2 If severely aff ected consider exchange blood transfusion. Further reading Prchal, J.T., et al. (2005) Red cell enzymes. Hematology Am Soc Hematol Edu c Program, 19 \200\22323. 96 CHAPTER 2 Red cell disorders Microangiopathic haemolytic anemia De \201 nition i RBC destruction caused when mechanical forces disrupt the physical RBC me mbrane integrity. Caused by trauma or vascular endothelial abnormalities. Causes \200 TTP/HUS \200\224E see Haemolytic uraemic syndrome, p.540; Thrombotic thrombocytopenic purpura, p.538. PET/HELLP (haemolysis, elevated liver enzymes, and low platelets). \200 Malignant tumor c irculations. \200 Renal abnormalities, e.g. acute glomerulonephritis, transplant rejection, malignant hypertension. Vasculitides, e.g. Wegener \200\231s, PAN, SLE. \200 DIC. \200 Anticancer and other dru gs, e.g. mitomycin C, tacrolimus, cyclosporin, ticlopidine, cocaine Cardiac \200\224prosthetic heart valves, grafts or patches, aortic stenosis and regurgitant jets. \200 March haemoglobinuria. \200 A-V malformations, e.g. Kasabach \200\223Merritt syndrome, A-V shunts. \200 Burns. Clinical \200 Varying degree of anemia \200\224most severe in DIC, TTP/HUS, and HELLP. Often associated with d platelets. \200 Blood fi lm shows marked RBC fragmentation, stoma tocytes, and spherocytes. Reticulocytosis often very marked. \200 Signs of underlying disease should be sought. Treatment \200 Diagnose and treat underlying disease. \200 Give folic acid and Fe suppl ements if defi cient. \200 TTP/HUS may require plasma exchange using FFP. Further reading Antman, K.H. et al. (1979). Microangiopathic hemolytic anemia and cancer: a review. Medicine (Baltimore), 58, 377 \200\22384. George, J.N. (2000). How I treat patient s with thrombotic thrombocytopenic purpura- hemolytic uremic syndrome. Blood, 96, 1223 \200 \2239. Moake, J.L. (2002) Thrombotic microangiopathies. N Engl J Med, 347, 589 \200\223600. 97 ACANTHOCYTOSIS Acanthocytosis Abnormal RBC shape (thorn-like surface protrusions) seen in a number of cond itions, inherited or acquired, aff ecting RBC membrane lipid structure. RBCs develop normal ly in marrow but once in plasma adopt characteristic shape. RBCs lose membrane and become p rogressively less elastic. Inherited conditions resulting in signi \201 cant acanthocytosis \200 A- -lipoproteinae mia. \200 McLeod phenotype (lacking Kell antigen). \200 In(Lu) phenotype. \200 In ass ociation with abnormalities of band 3 protein. \200 Hereditary hypo- -lipoproteinaemia. Acquired conditions resulting in signi \201 cant acanthocytosis \200 Severe liver diseas e. \200 Myelodysplastic syndromes. \200 Neonatal vitamin E defi ciency. Inherited conditions resulting in mild acanthocytosis \200 McLeod phenotype heterozygote. \200 PK defi ciency. Acquired conditions resulting in mild acanthocytosis \200 Post-splenectomy and hypospleni c states. \200 Starvation including anorexia nervosa. \200 Hypothyroidism. \200 Panhy popituitarism. A- \235\233\203-lipoproteinaemia Autosomal recessive. Congenital absence of apolipoprotein. i cholesterol:phospholipid ratio. RBC precursors normal. Usually obvious in early life with associated malabsorption of fat (including vitamins A, D, E, and K). Sp hingomyelin accumulates. Hematological abnormalities \200 Mild haemolytic anemia. \200 50 \200\22390% circulatin g RBCs are acanthocytic. \200 Reticulocytes mildly i. McLeod phenotype \200 d expression of Kell antigen on RBC. \200 Mild (compensated) haem olytic anemia. \200 10 \200\22385% acanthocytic RBCs in peripheral blood. Further reading Cooper, R.A. (1980). Hemolytic syndromes and red cell membrane abnormalitie s in liver disease. Semin Hematol, 17, 103 \200\22312. McBride, J.A., et al. (1970). Abnorm al kinetics of red cell membrane cholesterol in acanthocytes: studies in genetic and experi mental abetalipoproteinaemia and in spur cell anemia. Br J Haematol, 18, 383 \200\22397. 98 CHAPTER 2 Red cell disorders Autoimmune haemolytic anemia RBCs react with autoantibody complement l premature destruc tion of RBCs by reticuloendothelial system. Mechanism RBCs opsonized by IgG, recognized by Fc receptors on RES macrophages l phagocytos is. If phagocytosis incomplete remaining portion of RBC continues to circulate as spherocyt e (note: phagocytosis usually complete if complement involved). Seen in: \200 Haemolytic blood transfusion reactions. \200 Autoimmune haemolytic anemia (see Table 2.13). \200 Drug-induced haemolysis (some). Table 2.13 Types of autoimmune haemolyte anemia. Idiopathic Warm antibody induced 2 to lymphoproliferative disease, e.g. CLL, NHL 2 to other autoimmune diseases, e.g. SLE Idiopathic Cold antibody induced Cold haemagglutinin disease (CHAD) 2 to Mycoplasma infection Infectious mononucleosis Lymphoma Idiopathic Paroxysmal cold haemoglobinuria 2 to viral infection Congenital or tertiary syphilis Warm antibody induced haemolysis Extravascular RBC destruction by RES mediated by warm-reac ting anti- body. Most cases are idiopathic with no underlying pathology, but may be 2 to lymphoid malignancies, e.g. CLL, or autoimmune disease such as SLE. Epidemiology Aff ects predominantly individuals >50 years of age. Clinical features \200 Highly variable symptoms, asymptomatic or severely anemic. \200 Chronic compensated haemolysis. \200 Mild jaundice common. \200 Splenomegaly usual. 99 AUTOIMMUNE HAEMOLYTIC anemia Diagnosis \200 Anemia. \200 Spherocytes on peripheral blood fi lm. \200 ii reticulocy tes. \200 Neutrophilia common. \200 RBC coated with IgG, complement, or both (detect us ing DAT). \200 Autoantibody \200\224often pan-reacting but specifi city in 10 \200\22315% (Rh, mainly anti-e, anti-D, or anti-c). i LDH. \200 d serum haptoglobin. \200 Consider underlying lymphoma (BM, blood, and marr ow cell markers). Autoimmune profi le \200\224to exclude SLE or other connective tissue disorder. Treatment Prednisolone 1mg/kg/d PO tailing off after response noted (usually 1 \200\2232 we eks). If no response consider immunosuppression, e.g. azathioprine or cyclophosphamide. Spl enectomy should be considered in selected cases. IVIg (0.4g/kg/d for 5d) useful in refracto ry cases, or where rapid response required. Rituximab (anti-CD20) is emerging as a useful a gent for a range of refractory autoimmune disorders, including AIHA. Regular folic acid (5m g/d) is advised. Further reading Gehrs, B.C. and Friedberg, R.C. (2002). Autoimmune hemolytic anemia. Am J H ematol, 69, 258 \200\22371. Zecca, M. et al. (2003). Rituximab for the treatment of refract ory autoimmune hemolytic anemia in children. Blood, 101, 3857 \200\22361. 100 CHAPTER 2 Red cell disorders Cold haemagglutinin disease Describes syndrome associated with acrocyanosis in cold weather due to RBC agglutinates in blood vessels of skin. Caused by RBC antibody that reacts most strongly at temperatures <32 C. Complement is activated l RBC lysis l haemoglobinaemia and haemoglobinuria. May be idiopathic (1 ) or 2 to infection with Mycoplasma or EBV (infec tious mononucleosis). Clinical features \200 Elderly. \200 Acrocyanosis (blue discoloration of extremities, e .g. fi ngers, toes) in cold conditions. Chronic compensated haemolysis. \200 Splenomegaly usual. Diagnosis \200 Anemia. \200 ii reticulocytes. \200 Neutrophilia common. \200 +ve DA T \200\224C3 only. \200 Autoantibodies \200\224IgG or IgM: Monoclonal in NHL. \200 Polyclonal in infection-related CHAD. IgM antibodies react best at 4 C (thermal amplitude 4 \200\22332 C). \200 Specifi city: Anti-I (Mycoplasma). \200 Anti-i (infectious mononucleosis) \200\224causes little haemoly sis in adults since RBCs have little anti-i (cf. newborn i >>I). i LDH. \200 d serum haptoglobin. \200 Exclude underlying lymphoma (BM, blood, and marro w cell markers). \200 Autoimmune profi le to exclude SLE or other connective tissue disorder. Treatment \200 Keep warm. \200 Corticosteroids generally of little value. \200 Chlora mbucil or cyclophosphamide (greatest value when there is underlying B-cell lymphoma, occasi onally helpful in 1 CHAD). Plasma exchange may help in some cases. \200 If blood transfusion required use in-line bl ood warmer. \200 Splenectomy occasionally useful (note: liver is main site of RBC sequestration of C3b-coated RBCs) \200 Infectious CHAD generally self-limiting. Natural history Prolonged survival, spontaneous remissions not unusual, with periodic relap ses. 101 LEUCOERYTHROBLASTIC anemia Leucoerythroblastic anemia De \201 nition A form of anemia characterized by the presence of immature white and RBCs i n the peripheral blood. Mature white cells and platelets are also often reduced. Causes Marrow in \201 ltration by: \200 2 malignancy: commonly breast, lung, prostate, thyroid, kidney, colon. Myelofi brosis (a 1 myeloproliferative disorder, E see Primary myelofi brosis, p.292). Other hematological malignancy, e.g. myeloma and Hodgkin disease. \200 Rarely, severe hae molytic or megaloblastic anemia. Marrow stimulation by: \200 Infection, infl ammation, hypoxia, trauma (common in ITU pati ents). \200 Massive blood loss. When due to marrow infi ltration, there is often associated neutropenia thrombocytopenia . In cases with marrow stimulation there is often neutrophilia and thrombocytosis. Investigations \200 FBC and blood fi lm. Typical fi lm appearances are of i polychromasia due to reticulocytosis, nucleated RBCs, poikilocytosis (tear drop forms common in infi ltr ative causes), myelocytes and band forms, occasionally even promyelocytes and blast cells. Clotting screen \200\224where cause is 2 malignancy or infective, DIC may occur. BM is usually diagnostic \200 Hypercellular BM with normal cell maturation, typical of ma rrow stimulation causes. Infi ltration with neoplastic cells of a 2 malignancy may be identifi ed as abnormal clumps with characteristic morphology \200\224 immunohistochemistry may identify the 1 source, e.g. PSA for prostate. Increase in reticulin fi bres running in parallel bundles identifi es fi brotic infi ltrati ve cause \200\224usually myelofi brosis, but may occur with other hematological malignancy. Treatment \200 Diagnose and treat underlying cause if possible. \200 Supportive transfu sions as required, management of BM failure. Further reading Oster, W. et al. (1990). Erythropoietin for the treatment of anemia of mali gnancy associated with neoplastic bone marrow infi ltration. J Clin Oncol, 8, 956 \200\2236 2. 102 CHAPTER 2 Red cell disorders Aplastic anemia De \201 nition A gross reduction or absence of haemopoietic precursors in all 3 cell linea ges in BM resulting in pancytopenia in peripheral blood. Although this encompasses all situ ations in which there is myelosuppression, the term is generally used to describe those in which spontaneous marrow recovery is unusual. Incidence Rare 75 cases per million population annually. Wide age range, slight increase ar ound age 25 years and >65 years. 10 \227 more common in Asian people. Causes Divided into categories where aplasia is regarded as: \200 Inevitable: TBI dose of >1.5Gy (note: >8Gy always fatal in absence of graft rescue). Chemotherapy, e.g. high-dose busulfan. Hereditary: Fanconi syndrome \200\224stem cell repair defect resulting in abnormalities of skin, facies, musculo-skeletal system, and urogenital systems. BM failure often delayed until adulthood. Idiosyncratic: Chronic benzene exposure. \200 Drug-induced, but not dose related \200\224mainly gold, chloramphenicol, phenylbutazone, NSAIDs, carbamazepine, phenytoin, mesalazine. Genetic predisposition demonstrated for chloramphenicol. Post-viral: Parvoviral infections \200\224classically red cell aplasia but may be all elements. Devasta ting in conjunction with chronic haemolytic anemia, e.g. aplastic sickle crisis. Hepatitis viruses A, B, and C, CMV and EBV. Idiopathic: Constitute the majority of cases. Classi \201 cation \200 According to severity most clinically useful (see Table 2.14). \200 Defi nes highest risk groups. Table 2.14 Classifi cation of severity in aplastic anemia 2 of the following: Neutrophils Severe <0.5 \227 109/L <20 \227 109/L Platelets Reticulocytes <1% <0.2 \227 109/L and infection present Neutrophils Very severe 103 APLASTIC anemia Clinical features Refl ects the pancytopenia. Bleeding from mucosal sites common, with purp ura, ecchymoses. Infections, particularly upper and lower res- piratory tracts, skin, mouth , peri-anal. Bacterial and fungal infections common. Anemic symptoms usually less severe du e to chronic onset. Diagnosis and investigation \200 FBC and blood fi lm show pancytopenia, MCV may be i, fi lm morphology unremarkable. \200 Reticulocytes usually absent. \200 BM aspirate and trephi ne show gross reduction in all haemopoietic tissue replaced by fat spaces \200\224important to exclude hypocellular MDS or leukemia \200\224the main diff erential diagnoses. Flow cytometry using anti-CD55 and anti-CD59 will show lack of both membrane proteins. Ham \200\231s acid lysis test is now largely obsolete. Specialized cytogenetics on blood to exclude Fanconi syndrome (E see Fanconi \200\231s anemia, p.594). Complications \200 Progression to more severe disease. \200 Evolution to PNH \200\224oc curs in 7%. \200 Transformation to acute leukemia occurs in 5 \200\22310%. Treatment \200 Mild cases need careful observation only. More severe will need supportive treatment with red cell and platelet transfusions and antibiotics as needed. Blood product s should be CMV \210\222ve, and preferably leucodepleted to reduce risk of sensitization. Specifi c treatment options are between allogeneic transplant and immunosuppression. Sibling allogeneic transplant treatment of choice for those <50 years with sibling donor. Should go straight to transplant avoiding immunosuppression and b lood products if possible. Matched unrelated donor transplant should be considered in <25 years age group. Immunosuppressive options include antilymphocyte globulin (ALG) ciclosporin. Response to ALG may take 3 months. Refractory or relapsing patients may respond to a second course of ALG from another animal. Ciclosporin post-ALG looks promising. \200 Androgens or danazol may be useful in some cas es. Further reading Abkowitz, J.L. (2001). Aplastic anemia: which treatment? Ann Intern Med, 13 5, 524 \200\2236. Young, N.S. and Barrett, A.J. (1995). The treatment of severe acquired ap lastic anemia. Blood, 85, 3367 \200\22377. Young, N.S. et al. (1997). The pathophysiology o f acquired aplastic anemia. N Engl J Med, 336, 1365 \200\22372. 104 CHAPTER 2 Red cell disorders Paroxysmal nocturnal haemoglobinuria De \201 nition Rare acquired clonal abnormality of the haemopoietic stem cell leading to c hronic haemolysis, i risk of VTE and BM suppression rendering them more sensitive to comple ment-mediated lysis, most noticeable in RBCs. Cells lack phosphatidylinositol glycoproteins (PIG) transmem- brane anchors. Caused by mutation in X-linked PIG-A gene. Incidence Rare. Aplastic anemia is closely related. Clinical features \200 Chronic intravascular haemolytic anemia particularly overnight (?d ue to lower blood pH). Infections trigger acceleration of haemolysis. WBC and platelet production also often d. \200 Chronic haemolysis may induce nephropathy. \200 Haemoglobinuria usually results in Fe defi ciency. \200 ii tendency to venous thr ombosis particularly at atypical sites, e.g. hepatic vein (Budd \200\223Chiari syndrome), s agittal sinus thrombosis. Fatigue, dysphagia, and impotence occasionally seen. \200 BM failure often accompanies PN H. Diagnosis and treatment \200 FBC, blood fi lm \200\224polychromasia and reticulocytosis ( cf. AA). \200 BM aspirate and trephine biopsy \200\224usually hypoplastic with i fat spac e but with erythropoietic nests or islands distinct from AA. Ham \200\231s test (acidifi ed serum lysis) is invariably +ve though non-specifi c and seldom used now. Cellular immunophenotype shows altered PIG proteins, CD55, and CD59. Urinary haemosiderin +ve. Complications \200 May progress to more severe aplasia. \200 Transforms to acute leukem ia in 5%. \200 Serious thromboses in up to 20%. Treatment \200 Chronic disease \200\224supportive care may be satisfactory in mild cases. \200 Fe replacement usually required. \200 Trial of steroid/androgens/danazol may d sy mptoms and transfusion need. ALG/ciclosporin may be indicated for more severe cases as for aplastic anemia. Acute major thromboses should be treated aggressively with urgent thrombolysis and 10d of h eparin. Long-term warfarin mandatory. Consider warfarin prophylaxis after any one clotting episode. Severe cases <50 years should be considered for sibling allogeneic transplant if they have a donor \200\224consider MUD in <25 years age group if no sibling donor. 105 PAROXYSMAL NOCTURNAL HAEMOGLOBINURIA Eculizumab \200\224humanized monoclonal antibody against complement C5. Inhibits terminal c omplement activation. Recent TRIUMPH study showed that eculizumab able to stabilize Hb and d transfusion requirements in classical PNH. i risk of neisserial infection and patients re quire vaccination against Neisseria meningitidis 2 weeks before receiving study drug. Prognosis Median survival from diagnosis is 9 years. Major cause of mortality is thrombosis and marrow failure. Molecular genetic basis now estab- lished \200\224could be a candidate disease for gene transplantation. Further reading Hillmen, P. et al. (2004). Eff ect of eculizumab on hemolysis and transfusi on requirements in patients with paroxysmal nocturnal hemoglobinuria. N Engl J Med, 350, 55 2 \200\2239. Hillmen, P. et al. (1995). Natural history of paroxysmal nocturnal hemoglobinu ria. N Engl J Med, 333, 1253 \200\2238. Moyo, V.M., et al. (2004). Natural history of parox ysmal nocturnal haemoglobinuria using modern diagnostic assays. Br J Haematol, 126, 133 \200\2238. Parker, C. et al. (2005). Diagnosis and management of paroxysmal nocturnal haemo globinuria. Blood, 106, 3699 \200\223709. Rosse, W.F. (1997). Paroxysmal nocturnal hemoglob inuria as a molecular disease. Medicine (Baltimore), 76, 63 \200\22393. 106 CHAPTER 2 Red cell disorders Pure red cell aplasia De \201 nition A group of disorders characterized by reticulocytes <1% in PB, <0.5% mature erythroblasts in BM but with normal WBC and platelets. Incidence rare. May be caused by ab normal stem cells or 2 to auto- immune disease, viral infection, or chemicals. Congenital and acquired forms exist (see Table 2.15). Table 2.15 Classifi cation of red cell aplasia Congenital Diamond \200\223Blackfan anemia (DBA), E see Congenital red cell aplasia, p.590 Childhood: transient erythroblastopenia of childhood (TEC) Acquired Adults: 1 autoimmune or idiopathic 2 chronic: thymoma, hematological malignancies especially CLL, pernicious anemia, some so lid tumors, SLE, RA, malnutrition with ribofl avin defi ciency 2 transient: infections especially parvovirus B19, CMV, HIV, many drugs e.g. chlorampheni col, gold, dapsone, rifampin, sulfasalazine Recent interest following red cell aplasia in renal patients treated with SC EPO Clinical features \200 Lethargy usually only symptom of the anemia since slow onset. \200 No abnormal physical signs except of any underlying disease. Diagnosis and investigations \200 FBC shows severe normochromic, normocytic anemia with reticulocytes <1%. WBC and platelets n. BM shows absence of erythroblasts but is normocellular (distinguishes from aplastic anemia). Treatment \200 Treat underlying cause fi rst if identifi ed. \200 Remove thymoma. \200 If due to parvovirus B19, try IVIg (if congenital haemolytic anemia transfusion may be required). Assume immune origin if no other cause found and give prednisolone 60mg od PO as starter do se 740% response. Failure of response, try ciclosporin or ALG or azathioprine. Prognosis \200 15% spontaneous remission. 65% will respond to immunosuppression. 50% will relapse but 80% of relapsers will respond again. \200 A few progress to AA or AM L. Further reading Raghavachar, A. (1990). Pure red cell aplasia: review of treatment and prop osal for a treatment strategy. Blut, 61, 47 \200\22351. 107 IRON OVERLOAD Iron (Fe) overload Fe is an essential metal but overload occurs when intake of Fe exceeds r equirements and occurs due to the absence in humans of a physiolog- ical mechanism to excre te excess Fe. Sustained i Fe intake (dietary or parenteral) may result in Fe accumulation, overload, and potentially fatal tissue damage. Fe absorption is regulated at the enterocyte level by HFE and other proteins. Timing and pattern of tissue damage is determined by rate of accumula- tion, the quantity o f total body Fe and distribution of Fe between reticu- loendothelial (RE) storage sites and vulnerable parenchymal tissue. Fe accumulation in parenchymal cells of the liver, heart, p ancreas, and other organs is the major determinant of clinical sequelae. Haemochromatosis \200 Inherited (autosomal recessive) occurring in up to 0.5% population (N Europe). 10 \200\22313% N European Caucasians are heterozygotes. \200 Haemochromatosis locus is ti ghtly linked to the HLA locus on chromosome 6p and up to 10% of the population are heterozygous. \200 Single missense muta tion found in the homozygous state in 80% of patients. The gene designated HFE is an MHC class Ib gene. \200 2 mutations in HFE cause most cases of HH. C282Y is the most important. Most HH patients have this mutation (individuals homoz ygous for C282Y account for 790% cases of HH). Heterozygotes do not develop Fe overload pro viding there are no additional risk factors (e.g. alcohol, hepatitis). The second is H63D; this is less penetrant. A small proportion develop Fe overload. Compound heterozygotes (C28 2Y/H63D) may develop mild Fe overload. A third (uncommon) mutation is S65C. \200 Homozygo tes develop symptomatic Fe overload. Caused by failure to regulate Fe absorption from bowel causing progres- sive increase in to tal body Fe. Parenchymal accumulation occurs initially in liver then pancreas, heart, skin, and other organs rather than RE sites. Symptoms do not usually develop until middle age wh en body Fe stores of \211 15 \200\22320g have accumulated. Environmental factors (e.g. alc ohol use in males and menstruation in females) aff ect rate of accumulation and age at pres entation. Clinical expression of haemochromatosis is seen 10 \227 more commonly in \231 \202. Only 25% of heterozygotes show evidence of minor increases in Fe stores and clinical problems do not occur. Clinical manifestations of Fe overload only occur in homozygotes and presentation as \200 \230bronze diabetes \200\231 is characteristic (E see Table 2.16). Evaluation of Fe status \200 Indirect measure of Fe stores is serum ferritin estimation ( useful surrogate marker of total body Fe stores). Rises to maximum concentration of 4000mcg /L and may underestimate extent of Fe overload in some patients. Note: may be spuriously i by infection, infl ammation, or neoplasia. 108 CHAPTER 2 Red cell disorders Table 2.16 Clinical features of Fe overload (homozygous haemochromatosis) Skin pigmentation \200 Hepatic dysfunction Slate gray or bronze discolouration \200 Hepatomegaly, chronic hepatitis, fi brosis, \200 cirrhosis, hepatocellular carcinoma (20 \200\22330%) \200 Retinopathy, nephropathy, neuropathy, vascular Diabetes mellitus complications Gonadal dysfunction \200 Other endocrine dysfunction \200 Abdominal pain \200 Cardiac dysfunction Hypogonadism, impotence \200 Hypothyroidism, hypoparathyroidism, adrenal insuffi ciency Unknown etiology (25%) \200 Cardiomyopathy, heart failure, dysrhythmias (10 \200\22315%) \200 Arthropathy Chondrocalcinosis The % transferrin saturation provides confi rmatory evidence but no measure of the extent o f Fe overload. Useful screening test. If >60% in \231\202 or 45 \200\22350% in \231\200 r epeat and investigate further. MRI useful tool for assessment of Fe overload in heart (T2*) and liver (T2*/R2). Useful for monitoring response to treatment (e.g., venesection). Liver biopsy provides a direct albeit invasive measure of Fe stores (% Fe concentration by weight) and visual assessment of Fe distribution, and the extent of tissue damage. Only ind icated if histological investigation also planned, e.g., staging of cirrhosis. MRI T2* useful tool for assessment of Fe overload in heart, liver and other sites. SQUID (superconducting quantum interference device) uses high- power magnetic fi eld and de tectors that measure the interference of Fe within the fi eld. Linear correlations have bee n demonstrated between SQUID measurements and liver biopsy Fe levels but restricted availab ility. Diagnosis May be diffi cult to diff erentiate haemochromatosis from Fe overload 2 to othe r causes, particularly that associated with chronic liver disease. Recent identifi cation o f HFE gene will provide a tool for more defi nitive diagnosis and screening of relatives (p reviously performed by serum fer- ritin estimation). Management \200 Aim to reduce ferritin to <50mcg/L and prevent complications of overload. Achieved by regular venesection (500mL blood = 200 \200\223250mg Fe) on weekly basis until Fe defi ciency develops (may take many months). 109 IRON OVERLOAD Hb should be measured prior to each venesection and response to therapy can be monitored by intermittent measurement of the serum ferritin. R2 liver (some individuals also require T2* heart) 6 \200\22312-monthly. \200 Once Fe def i ciency develops a maintenance regimen can be commenced with venesection every 3 \200\2234 months. Avoid alcohol, exogenous Fe. \200 Liver cirrhosis and hepatoma risk are not reversible. Natural history Cirrhosis and hepatocellular carcinoma are the most common causes of death in patients with haemochromatosis and are due to hepatic Fe accumulation. Cirrhosis does no t usually develop until the hepatic Fe concentration reaches 4000 \200\2235000mcg/g of live r (normal 50 \200\223500mcg/ g). Hepatocellular carcinoma is the cause of death in 20 \200 \22330% but does not occur in the absence of cirrhosis which increases the risk over 200 \227. If venesection can be commenced prior to the development of cir- rhosis and other com plications of haemosiderosis the life expectancy is that of a normal individual. Reduction of Fe overload by venesection has only a small eff ect on symptomatology which has already developed: skin pigmentation diminishes, liver function may improve, cardiac abnor- malitie s may resolve, diabetes and other endocrine abnormalities may improve slightly, arthropathy is unaff ected. Further reading Beutler, E. et al. (2002). Penetrance of 845G >A (C282Y) HFE hereditary ha emochromatosis mutation in the USA. Lancet, 359, 211 \200\22318. Bulaj, Z.J. et al. (2000). Disease-related conditions in relatives of patients with hemochromatosis. N Engl J Med, 34 3, 1529 \200\22335. Olynyk, J.K. et al. (1999). A population-based study of the clinical ex pression of the hemochromatosis gene. N Engl J Med, 341, 718 \200\22324. Sanchez, A.M. et a l. (2001). Prevalence, donation practices, and risk assessment of blood donors with hemochr omatosis. JAMA, 286, 1475 \200\22381. 110 CHAPTER 2 Red cell disorders Transfusion haemosiderosis Fe overload occurs in patients with transfusion dependent anemia , notably thalassaemia major, Diamond \200\223Blackfan syndrome, aplastic anemia, and acqui red refractory anemia. Also increasingly seen in transfused sickle patients. In many of the se conditions Fe overload is aggravated by physiological mechanisms which promote i dietary absorption of Fe in response to ineff ective erythropoiesis. Each unit of blood contains 2 00 \200\223250mg Fe and average transfusion dependent adult receives 6 \200\22310g of Fe/ye ar. Distribution of Fe is similar to haemochro- matosis with primarily liver parenchymal ce ll accumulation followed by pancreas, heart, and other organs. Cardiac deposition occurs in patients who have received 100 units of blood (20g Fe) without chelation, and is followed by damage to the liver, pancreas, and endocrine glands. Pattern of distribution also determ ined by genetic factors. Clinical features of Fe overload in children who require transfusion support for hereditary anemia are listed. Similar problems excluding those related to growth and sexual maturatio n develop in patients who commence a transfusion program for acquired refractory anemia in later life. Features of transfusion haemosiderosis in hereditary anemia \200 Growth retardation in 2n d decade. \200 Hypogonadism \200\224delayed or absent sexual maturation. \200 Skin pigm entation \200\224slate gray or bronze discolouration. \200 Hepatic dysfunction \200\224he patomegaly, chronic hepatitis, fi brosis, cirrhosis, hepatocellular carcinoma. Diabetes mellitus. \200 Other endocrine dysfunction \200\224rarely hypothyroidism, hypoparathyroidism, adrenal insuffi ciency. Cardiac dysfunction \200\224cardiomyopathy, heart failure, dysrhythmias (main cause of death). Death from heart disease in adolescence. Management \200 Fe chelation therapy using currently available chelation agents (desferrioxamine, deferiprone and deferasirox) is the mainstay of treatment for patients wi th transfusion haemosiderosis, who remain anemic. Haemosiderosis due to previous transfusio ns in conditions where Hb now normal, e.g. treated AML, may be venesected to remove Fe. Regular treatment is required in transfusion dependent children if they are to avoid the consequences of Fe overload in the 2nd decade of life. SC administration of desferrioxamine by portable syringe pump over 9 \200\22312h on 5 \200\2237 nights/week is a common regimen. Ascorbic acid supplementation is given at the same time to aid excretion. 111 TRANSFUSION HAEMOSIDEROSIS Deferiprone (Ferriprox) is orally active but needs to be taken 3 times a day. Can also be g iven as combination therapy with desferrioxamine. Contraindicated for use in patients with DBA. Deferasirox (Exjade) is the newest licensed agent. Given as a suspension once daily. Avoid in renal impairment. Chelation therapy is started after a year of regular transfusions or when ferritin > 1000mc g/L. d hepatic Fe and improves hepatic function, promotes growth and sexual development, an d protects against heart disease and early death. Newer chelation therapies are under investigation. Natural history The prognosis of the underlying hematological condition in transfusion- dep endent elderly patients may eliminate the need for Fe chelation. In others with a longer li fe expectancy, a period of venesections (if good Hb) or iron chelation therapy should be co nsidered.