SYNOPSIS
Rajiv Gandhi University of Health Sciences, Karnataka,
Bangalore
Name of the candidate : Dr. SALMAN AKTHER
Guide : Dr. JAYAPRAKASH C.S.
Topic : Study of Erythrocyte and
Reticulocyte indices in Iron
Deficiency Status
Course and Subject : M.D.PATHOLOGY
Department of Pathology,
Father Muller Medical College,
Kankanady, Mangalore – 575002.
July 2013
RAJIV GANDHI UNIVERSITY OF HEALTH SCIENCES
BANGALORE, KARNATAKA
PROFORMA FOR REGISTRATION OF THE SUBJECT FOR
DISSERTATION
1. / Name of the candidate and address (in block letters) / DR. SALMAN AKTHERP.G. RESIDENT
FATHER MULLER MEDICAL COLLEGE, KANKANADY,
MANGALORE – 575002.
2. / Name of the institution / FATHER MULLER MEDICAL COLLEGE, KANKANADY,
MANGALORE – 575002
3. / Course of study and subject / M.D. PATHOLOGY
4. / Date of admission to the course / 31-07-2013
5. / Title :
STUDY OF ERYTHROCYTE AND RETICULOCYTE INDICES IN IRONDEFICIENCY STATUS
6. / BRIEF RESUME OF THE INTENDED WORK:
6.1 NEED FOR THE STUDY
Iron status reflects balance between rate of erythropoiesis and iron stores in the body. Iron deficiency is one of the most common nutritional deficiency in world and a leading cause of anemia.(1)Anemia is very common and significant in country like India, Hence early diagnosis is of utmost importance .Anemia is defined as decreased concentration of Hemoglobin(Hb) in the blood, cutoff value depending on age and gender. In practice Anemia is usually diagnosed based on reduction of Hematocrit and Hb concentration, but isolated Hb measurement has both low specificity and low sensitivity. Several studies in the field of iron metabolism and erythropoiesis are modifying the traditional concepts on anemia.
These studies point out the need for reliable diagnostic parameters that allow better evaluation of the causes underlying apparently similar clinical conditions. Thus implying stronger collaboration between laboratory professionals and clinicians in order to optimize early diagnosis of anemia by new analytical parameters viz. Rsf (Red cell size factor) and LHD% (Low hemoglobin density percentage).Not enough work has been done on these parameters and such a study will be very much helpful in the future.
6.2 Review of literature
Iron homeostasis acts at both the cellular and the systemic levels. It involves a complex system of different cell types, transporters, and signals. To maintain systemic iron homeostasis, communication between cells that absorb iron from the diet (duodenal enterocytes), consume iron(mainly erythroid precursors), and store iron (hepatocytes and tissue macrophages) must be tightly regulated.(2)
Functional iron deficiency is defined as an imbalance between the iron needs for erythropoiesis and the iron supply, with the latter not maintained at sufficient rate for adequate hemoglobinization.(3) In iron deficiency anemia (IDA) iron supply depends on the quantity of iron stored in the body, while in functional iron deficiency(iron restricted erythropoiesis) supply depends on the rate of mobilization of iron from the stores.
Serum ferritin which is an indicator of iron storage but not of iron supply, is a positive acute phase reactant; while transferrin is a negative acute phase reactant.(4-6)
For these reasons, an iron deficient erythropoiesis may occur despite normal serum ferritin and transferrin saturation values.(7)
Therefore novel parameters like Red cell size factor (Rsf), low hemoglobin density (LHD %), Reticulocyte hemoglobin content (CHr) and percentage of hypochromic cells (%Hypo) have been explored. Formulae to obtain these values are hereby given:
(1) Rsf = √[ MCV * MRV](8)
Red blood cell Size Factor (Rsf) joins together the volume of mature red cells (MCV) and the volume of reticulocytes (MRV), both related to erythropoietic activity and hemoglobinization. Reference range 91.1-114.4fl.(9)
(2) LHD% = 100*√ 1-[1/{1+eᶺ1.8(30-MCHC)}](4)
Low hemoglobin density (LHD %) derives from the traditional mean cell hemoglobin concentration (MCHC), using the mathematical sigmoid transformation. Reference range 0-4%(10)
(3) CHr (directly measured by Siemen's analyzer) is assessment of the incorporation of iron into erythrocyte hemoglobin (Hb) and thus an estimate of the recent functional availability of iron into the erythron.(11) CHr is a sensitive indicator of iron deficient erythropoiesis. (potential to be more accurate of iron status in Hemodialysis Patients)
(4) %Hypo (directly measured by Siemen's analyzer) is defined as the percentage of red blood cells with Hb concentration less than 280 g/L.(8) The measurement of %Hypo is a sensitive method for quantifying the hemoglobinization in mature red cells. Because of the long circulating life span of mature erythrocytes %Hypo values are related to iron status in the last 2-3 months, and have been recognized as an indicator of iron deficiency.(7)
Rsf and LHD% thus have been used as a diagnostic tool; together with biochemical markers and interest have been generated in the use of these two markers in particular available on the modern analyzers (Beckman-Coulter analyzers).
Clinical trends of hematological monitoring of the patients on frequent basis have created additional pressure on limited man power resources. Automated analyzers have substantially changed during the last20 years. Modern counters not only provide information about the reticulocyte counts but also about the characteristics of these cells (size or Hb content),related to the quality of the erythropoiesis, giving information of the current erythropoietic activity of the bone marrow.
Table 1: The new biomarkers of hypochromia now reported by the analyzers of different manufacturers.
RBC Parameter / Abbreviation (unit) / Company
Hypochromic / Hypo (%) / Siemens
Reticulocyte Hb content / CHr (pg) / Siemens
Hypochromic RBC % / HPO (%) / Abbott
Reticulocyte Hb content / MCHr (pg) / Abbott
Hypochromic RBC / Hypo He (%) / Sysmex
Reticulocyte Hb equivalent / Ret He (pg) / Sysmex
Low Hb density / LHD (%) / Beckman Coulter
Correct interpretation of the results requires correlation of the equipment knowledge with the clinical information.
Advantages:
(1) rapid and automated;
(2) rapid information to Clinicians about iron status;
(3) monitor response to therapy;
(4) Economical (simultaneous in course of routine blood counts).
Aids clinicians in
· assessing true status of iron by providing Hb content.
· detect functional iron deficiency
· considering other differential diagnosis
An accurate diagnosis depends on a Clinico-pathological Correlation nevertheless.
6.3 Objective of the study:
a) Relation between Rsf and LHD% in iron deficiency profile.
b) Significance of Rsf and LHD% in iron deficient status.
7. / MATERIALS AND METHODS
7.1 Source of data: The EDTA treated venous blood specimens received in hematology laboratory at Father Muller Medical College Hospital, department of Pathology will be included in this study.
7.2 Sample size: A minimum of 150 cases of peripheral smears showing Microcytic Hypochromic Anemia will be selected using purposive sampling technique based on inclusion exclusion criteria.
Design of the study: Correlative study, sampling technique is based on exclusion and inclusion criteria
Duration of the study: The study period will be from November 2013 to January 2015
7.3 METHOD OF COLLECTION OF DATA
Study will include patients of Father Muller Medical College Hospital, from November 2013 to January 2015.The data will be collected in the following manner
Clinical details comprising of
· Age
· Gender
After noting history and examination findings ,basic investigations will be performed and the following parameters are obtained
· Hemoglobin
· MCV
· MRV
· Mchc
· Mch
· Rsf
· LHD%
Duration of the study:
The study period will be from November 2013 to January 2015
Inclusion Criteria
Adult patients (age>18 yrs) with peripheral smears showing Microcytic Hypochromic Anemia will be included in the study (Hb < 10 g/dL)
Exclusion criteria
· Pediatric age group
· Normocytic Nornochromic anemia
· Macrocytic anemia
· Anemias with bleeding disorders
· Chronic anemias
Data Analysis
Collected data will be analysed by mean, standard deviation, Karl Pierrson correlation coefficient, t test.
7.4 Does the study require any investigations or interventions to be conducted on patients or other humans or animals
Yes. The study requires collection of blood samples for examination. Voluntary informed consent will be obtained from the subjects of the study
7.5 Has ethical clearance been obtained from your institution in case of 7.4?
Yes
INFORMED CONSENT FORM
Name of the volunteer:
Date:
Sex:
Age:
Address:
This is to declare that I ______have understood every aspect of the study “Study of Erythrocyte and reticulocyte indices in iron deficiency status” being conducted by Dr.Salman Akther, post graduate student of the Department of pathology, FMMC, Mangalore.
I voluntarily give my consent for the tests to be performed using my blood sample. The details of the study have been explained to me in an understandable manner and to my satisfaction. I have read this completely and I am sure that the data generated will be kept in at most confidence.
I completely agree with this and shall follow all medical directions given to me. By my own decision and self interest, I give my consent to take part in this study.
I keep my option to leave the study at any time I desire to do so. I am allowed to freely ask questions regarding the research study. I do agree to take part in the study voluntarily.
Signature of the volunteer:
Signature of the witness:
Signature of the principal investigator.
8. / LIST OF REFERENCES:
1. Thomas C, Thomas L. Biochemical Markers and Hematologic Indices in the Diagnosis of Functional Iron Deficiency. Clinical Chemistry.2002;48:1066-1076
2. Swinkela DW, Janssen MCH, Marx JJM. Hereditary hemochromatosis genetic complexity and new diagnostic approaches. Clinical Chemistry: 2006;52(6):950-968.
3. Cavill I, Macdougall IC. Functional iron deficiency. Blood.1993;82(4):137
4. Fishbane S, Imbriano LJ, Kowalski Ea, Maesaka JK. The evaluation of iron status in patients receiving recombinant human erythropoietin. Clin J Am Soc Nephrol.1996;7:654-657.
5. Mast A. The clinical utility of peripheral blood tests in the diagnosis of iron deficiency anemia. Bloodline.2001;1:7-9.
6. Coyne D.Iron indices: what do they really mean? Kidney Int Suppl.2006 May;69(101):S4-S8.
7. Urechaga E, Borque L, Escanero JF. Erythrocyte and Reticulocyte Indices on the LH 750 as Potential Markers of Functional Iron Deficiency. Anemia. 2010;2010:625919. doi: 10.1155/2010/625919.
8. Urechaga E, Borque L, Escanero JF. Biomarkers of Hypochromia: The Contemporary Assessment of Iron status and Erythropoiesis. Biomed Res Int. 2013;2013:603786. doi: 10.1155/2013/603786.
9. Urrechaga E. Clinical utility of the new Beckman-Coulter parameter red blood cell size factor in the study of erythropoiesis. Int J Lab Hematol. 2009 Dec:31(6):623-9. doi: 10.1111/j.1751-553X.2008.01088.x. Epub 2008 Sep 4
10. Urrechaga E. The new mature red cell parameter, low hemoglobin density of the Beckman-Coulter LH750:clinical utility in the diagnosis of iron deficiency. Int J Lab Hematol.2010 Feb;32 (1 Pt 1) e144-50. doi: 10.1111/j.1751-553X.2008.01127.x. Epub 2009 Feb 9.
11. Lowenstein L. The mammalian reticulocyte. Int Rev Cytol.1959;8:135-139.
9. / SIGNATURE OF THE CANDIDATE -
10. / REMARKS FROM GUIDE- / Rsf and LHD% are simple and reliable parameters, readily available in automated analyzers. Can be used for early diagnosis of anemia
11. / 11.1 NAME & DESIGNATION OF GUIDE / DR. JAYAPRAKASH C.S. MD
PROFESSOR
DEPARTMENT OF PATHOLOGY
FR. MULLER MEDICAL COLLEGE
MANGALORE – 575002
11.2 SIGNATURE
11.3 HEAD OF DEPARTMENT / DR. HILDA FERNANDES, MD
PROFESSOR AND HOD OF
PATHOLOGY
DEPARTMENT OF PATHOLOGY
FR. MULLER MEDICAL COLLEGE
11.4 SIGNATURE
12 / 12.1 REMARKS OF THE CHAIRMAN AND DEAN
12.2 SIGNATURE
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