IRON DEFICIENCY ANEMIA

includes breifs about the iron deficiency and in elaboration the life diagnosis of iron deficiency

1. IRON DEFICIENCY ANEMIA LAB DIAGNOSIS [email protected]

2. Contents Introduction Iron metabolism Causes of Iron deficiency Stages of IDA Clinical features of iron defeciency anemia Lab diagnosis of IDA Evaluation of anemia Treatment of anemia

3. INTRODUCTION Anemia is functionally defined as an insufficient RBC mass to adequately deliver oxygen to peripheral tissues. Iron deficiency anemia (IDA) is characterised by microcytic hypochromic red cells with MCV < 8O ft and MCH < 25 pg. PREVALENCE OF IDA Iron deficiency is the most widespread form of malnutrition affecting nearly two billion people the world over. It is the most common anemia prevalent in India. Prevalence of iron deficiency in Children in India varies from 35-45%. frequency is higher in females In pregnant females it is round 45%-60%.

4. IRON METABOLISM Iron is present in hemoglobin, myoglobin and iron containing enzymes of Cytochrome system. Iron is essential for DNA, RNA and protein synthesis. Every day about 30 mg iron is used to make new hemoglobin. • Daily iron loss is around 1 mg. • In women menstruation and childbirth increase iron losses to about 1.5 mg/day. The total content of iron in the body - about 3.5 to 4 g. > 75-80% belongs to the hemoglobin > 20 - 25% reserve > 5-10% part of the myoglobin > 1% is part of the enzyme for the tissue respiration

5. IRON ABSORPTION DIETARY IRON There are 2 types of iron in the diet; heme iron and non- heme iron. Heme iron is present in Hb containing animal food like meat, liver & spleen. Non- heme iron is obtained from cereals, vegetables & beans.

6. IRON ABSORPTION Site- Proximal small intestine i.e. duodenum (first part- maximum absorption) and jejunum. 10% of dietary iron is absorbed it is determined by intraluminal factor i.e. pH and redox potential. Therapeutic ferrous iron is well absorbed on empty stomach. Heme iron is not affected by ingestion of other food items. Heme iron → Acid and gastric juices release it from apoprotein → Oxidised → hemin → directly absorb through mucosal cells.

7. IRON ABSORPTION Steps involved in the absorption of Non Heme iron in intestine are :- reduction of ferric to ferrous ions apical uptake intracellular storage basolateral release.

8. Most body iron is present in hemoglobin in circulating red cells. The macrophages of the reticuloendotelial system store iron released from hemoglobin as ferritin and hemosiderin . In the plasma, total iron averages 110 μg / dL . Majority bound to the transferrin (capacity to bind 330 μg of iron per deciliter). So only one third of transferrin is saturated.

9. Iron absorption at molecular level : Iron is converted from Fe 3+ to Fe 2+ by ferrireductase (DCYTB). Fe2+ transported across mucosal surface of enterocyte by DMT1, stored as ferritin . Ferritin releases Fe2+ which is transported across basolateral surface of enterocyte with help of ferroportin . Fe 2+ converted back to Fe 3+ by Hephaestin . Fe 3+ binds to transferrin in plasma.

10. FACTORS AFFECTING IRON ABSORPTION: Enhancers: Ascorbic acid Citric acid Amino acid Sugars in diet Inhibitors: Tannate Carbonate Oxalate Phosphate

11. Causes of iron deficiency 4 major causes 1> Decreased intake 2> Decreased absorption 3> Increased demand 4> Chronic Blood loss

12. 1> Decreased intake Decreased iron in the diet – Vegetarian diet – Low socioeconomic status – Lack of balanced diet or poor intake 2>Decreased absorbtion – Gastric surgery – Achlorhydria – Duodenal pathology – Chronic renal failure patients – Coeliac Sprue 3 >Increased demands • Pregnancy • Lactation • Growing infants and children • Menstruating women • Multiparity • Parturition

13. 4> Increased iron loss Menorrhagia Gastrointestinal hemorrhage • P.Ulcer • Oesophagitis • Varices • Hiatal hernia • Malignancy • Angiodysplasia • Diverticulosis • Meckel diverticula • Colitis or imperforated bowel disease • Hemorrhoids • NSAID use • Parasites Bleeding disorder Pulmonary lesions with bleeding Hemoglobinuria – hemosiderinuria (chronic intravascular hemolysis ) Hemodialysis Hematuria (chronic) Frequent donation– 250 mg iron per unit-blood

14. Genetic causes

15. Clinical features Fatigue and Other Nonspecific Symptoms – irritability, palpitations, dizziness, breathlessness, headache, and fatigue Neuromuscular System – impair muscular performance, abnormalities in muscle metabolism , behavioral disturbances, – Neurologic development in infants and scholastic performance in older children may be impaired. – Sometimes neuralgia pains, vasomoto disturbances, or numbness and tingling.

16. Laboratory diagnosis Complete blood count RBC indices Peripheral blood smear Serum iron profile Bone marrow examination

17. IRON DEFICIENCY ANEMIA MCV - Reduced ( N : 80-100 fl) MCH - Reduced ( N : 27-32 pg) MCHC– Normal to reduced (N: 30-34 mg/dl) Iron- Reduced (N: 4 gm ) TIBC- Increased (N: 47-70 μmol /l) Transferin Saturation- Reduced (N :16-50%) Ferritin - Reduced (N:15–300 μ g/l) RDW: High ( N : 11.5- 14 %) Reticulocytes: Normal/Low (N: 0.5- 2.5%) Platelates : Normal/Low/High WBC: Normal/Low Smear: Hypochromia , microcytosis , anisopoikilocytosis may also present

18. BONE MARROW EXAMINATION Early stage - Normoblastic hyperplasia – Normoblasts - • smaller than normal • deficient in hemoglobin • Irregular shaped with frayed margins – polychromatic normoblasts - pyknotic nuclei, vacuolated cytoplasm and irregular in outline ( micronormoblastic erythropoiesis ) – absence of stainable iron

19. Marrow film- Iron deficiency anemia

20. Marrow film- prussian blue stain Normal Marrow Iron deficiency Anemia

21. Stages in the Development of Iron Deficiency Anemia Pre-latent – – reduction in iron stores without reduced serum iron levels – Hb , MCV, Transferrin saturation- Normal, Iron absorption -increase, Serum ferritin and marrow iron reduced – no clinical manifestation Latent- – iron stores are exhausted, but the blood hemoglobin level remains normal – index of the blood within the standard – clinical picture is caused by the sideropenic syndrome Iron Deficiency Anemia- – blood hemoglobin concentration falls below the lower limit of normal – the clinical manifestations in the form of sideropenic syndrome and general anemic symptoms

22. Serum Iron Reference interval is 50–160 μg / dL Determination of iron status requires- – Estimate of the amount of haemoglobin iron (usually by measuring the haemoglobin concentration ( Hb ) in the blood). – Level of storage iron (measuring serum ferritin concentration). Assay of serum iron- It is modification of method recommended by the International Council for Standardization in Haematology (ICSH) and is based on the development of a coloured complex when ferrous iron released by serum protein denaturation in the presence of reducing agent is treated with a chromogen solution. Alternate methods- 1. Microtitre plate method 2. Automated Methods for Serum Iron- non-precipitation method. INTERPRETATION Level reduced in - Iron deficiency anemia – Anemia of chronic diseases – Infections

23. Serum ferritin Normal value- 15–300 μg /L (Men>women) Water-soluble complex of iron hydroxide with the protein apoferritin . It is located in cells of the liver, spleen, bone marrow . Ferritin assay- Immunoradiometric assay MOA- Excess radiolabelled antibody react with ferritin and antibody not bound to ferritin was removed with an immunoadsorbent . Interpretation – • Ferritin is the basic protein which deposits iron. • Serum ferritin concentration reflects iron reserve of individual. • Reduced in iron deficiency anemia. • It is acute phase reactant so raised in some hepatocellular diseases, malignancies, and inflammatory diseases, so may give disproportionately high estimate of storage iron.

24. Transferrin • Transferrin has a very high affinity for iron at neutral pH and iron release takes place through a specific membrane receptor LIMITATION • The concentration of transferrin is subjected to the daily variations • Acute inflammation contributes to lowering the transferrin level CLINICAL SIGNIFICANCE • Basic clinical index for the differentiation between the iron-deficiency ([TF]↑) and hemolytic anemia ([TF]↓) • More precise index than total iron binding capacity • After the liberation of iron from the complex, TF ion of Fe3+ must be restored into Fe2+

25. Serum (Total) Iron-Binding Capacity (TIBC) Iron in plasma binds to transferrin and TIBC is the measure of this protein. The additional iron-binding capacity of transferrin is known as the unsaturated iron-binding capacity (UIBC). TIBC = UIBC + serum iron concentration . TIBC( μ mol/l) = Transferrin conc (gm/l) × 25 Estimation of TIBC- By adding an excess of iron to a solution and measuring the iron retained in solution after the addition of a suitable reagent such as ‘light’ magnesium carbonate or an ion-exchange resin that removes excess iron. Principle- Excess iron as ferric chloride is added to serum. Any iron that does not bind to transferrin is removed with excess magnesium carbonate. The iron concentration of the iron saturated serum is then measured. Interpretation-Raised in iron deficiency anemia

26. UIBC DETERMINATION The UIBC may be determined by methods that detect iron remaining and able to bind to chromogen , after adding a standard and excess amount of iron to the serum. The UIBC is the difference between the amount added and the amount binding to the chromogen . METHODS- – Chromogen solution – Microtitre tray UIBC is being evaluated as a screening test for iron overload in genetic haemochromatosis .

27. Serum Transferrin (Beta-globulin) Main function - transport of absorbed iron in the depot (liver, spleen), into the medullary erythroid predecessors and into the reticulocytes . Basic place of synthesis - liver. Reference interval for adults is 200–300 μg / dL (2.0–3.0 g/l ) 1 mg of transferrin binds 1.4 μg of iron. ESTIMATION OF SERUM TRANSFERRIN- by an immunological assay-Rate immunonephelometric methods INTERPRETATION • An increase in the content of transferrin with lowering in the level of iron of serum is characteristic for the iron- deficiency state. • A decrease in the level of transferrin can be with the damage of the liver (different genesis) and with the loss of protein (for example, in nephrotic syndrome). • The level of transferrin is increased in the last term of pregnancy.

28. Transferrin saturation The transferrin saturation is the ratio of the serum iron concentration and the TIBC expressed as a percentage Normally, this is 20%–55%. A transferrin saturation of <16% is usually considered to indicate an inadequate iron supply for erythropoiesis . Used for detection of genetic haemochromatosis . Normal diurnal variation serum iron is as much as 30% with highest values in the morning and lowest values late in the day. Fasting morning blood specimens are preferred for the diagnosis of iron deficiency. Transferrin Index- It is serum iron concentration ( μmol /l) divided by the transferrin concentration (determined immunologically and expressed as μ mol/l)

29. SERUM TRANSFERRIN RECEPTOR There are two types of transferrin receptors TfR1 and TfR2 . TfR1 is essential for tissue iron delivery Transferrin binds to TfR1, the complex is internalized and iron is released when the pH of the internal vesicles is reduced to about 5.5 .

30. Serum Transferrin Receptor–to– Serum Ferritin Ratio ( TfR /F) New approach to estimate total body iron stores. Have limited value in identifyin anemia of chronic disease (ACD) Better utilized in identifying iron defiiency anemia coexisting with ACD If the value comes greater than 1.5 : signifies iron deficiency If the value comes lower than 1.5 : signifies anemia of chronic disease.

31. DDs The differentials of microcytic hypochromic anemia are: Iron deficiency anemia Sideroblastic anemia Thalassemia Anemia of chronic disease Lead poisoning.

32. Treatment Deworming Oral Iron therapy

33. Parentral iron Erythropoeitin Packed cell volume

34. Response to treatment Feeling of well being Retic count start increasing within 5 to 7 days Iron stores replenishment in 3 to 6 months (after continuous therapy)

35. Bibliography Wintrobes clinical hematology 14 th edition Robbbins Basic pathology 10 th edition Atlas and text of hematology by Dr Tejinder singh 4 th edition Dacie and lewies Practical Hematology 12 th edition Essetials of hematology Shirish M Kawthalkar 2 nd edition www.slideshare.com Google images

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