THE ROLE OF CLINICAL BIOCHEMISTRY IN THE

DIAGNOSIS OF CRETINISM
BY

MLS. JOHN FAVOUR MANA

(INTERN)

DEPARTMENT OF CHEMICAL PATHOLOGY

FEDERAL MEDICAL CENTRE BIDA, NIGER STATE

SUPERVISED BY:
MLS ANIBASA ABDULAZEEZ
6th JULY, 2023 1
 OUTLINE
• INTRODUCTION
• EPIDEMIOLOGY
• PHYSIOLOGY
• AETIOLOGY
• PATHOPHYSIOLOGY
• SIGNS AND SYMPTOMS
• DIAGNOSIS
• TREATMENT
• CONCLUSION
• RECOMMENDATIONS
2
• REFERENCES
 LIST OF ABBREVIATIONS
• CH: Congenital Hypothyroidism
• CLIA: Chemiluminescence Immunoassay Analyzer
• CNS: Central Nervous System
• ELISA: Enzyme Linked Immunosorbent Assay
• ICP-MS: Inductively Coupled Plasma Mass Spectrometry
• NCH: Neonatal Congenital Hypothyroidism
• NNJ: Neonatal Jaundice
• PP: Polypropylene
• T4: Thyroxine
• T3: Triiodothyronine
• TGB: Thyroxine- Binding Globulin
• TRH: Thyrotropin Releasing Hormone
• TSH: Thyroid Stimulating Hormone
3
 INTRODUCTION 1/3

Iodine deficiency is a major public health problem worldwide,

especially in pregnant women and young children. Few among the
devasting outcomes of this deficiency in pregnancy are; perinatal
mortality, neonatal hypothyroidism, goiter, still births, abortions and
congenital anomalies (Jibril et al., 2016)

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 INTRODUCTION 2/3
• Cretinism (CH) is a condition of severe physical and mental
retardation due to iodine deficiency, specifically deficiency of thyroid
hormones during early pregnancy.
• Cretinism is the most serious iodine deficiency disorder and occurs
when a pregnant women is severely iodine deficient.
(Lucas et al., 2010)

5
 INTRODUCTION 3/3
• It is either due to endemic cretinism or due to sporadic cretinism.
• It is typically diagnosed during infancy or childhood. The congenital
hypothyroid child may appear normal at birth or present with slightly
increased head size due to myxedema of the brain.

(Byju, 2022)

6
 EPIDEMIOLOGY 1/2
The incidence of CH varies depending on the country, race and other
factors.
• It has been found to be as low as 1:3000 in America and as high as
1:420 live birth in Isfahan, Iran.
• In central Africa, as many as 10% of newborns may have both low
cord blood T4 concentration and TSH concentrations over 100 mlU/L
• The estimated prevalence of congenital hypothyroidism (CH) in
Nigeria is 42,500 per year. (Kayode-Adedeji &Alikah,
2015)
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 EPIDEMIOLOGY 2/2
• Available studies have shown that the incidence of NCH was 1.5% in
Saki, a town in Oyo state, Nigeria, 16.4% in Bassa ( a town with high
prevalence of endemic goitre in Northern Nigeria), and 3.8% in Jos.
• The aetiologies include: Thyroid dysgenesis (75%), Thyroid
dyshormonogenesis (10%), Transient hypothyroidism (10%) and
Hypothalamic-pituitary (TSH) deficiency (5%)

(Kayode-Adedeji &Alikah, 2015)

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 HYPOTHALAMUS PITUITARY-THYROID AXIS

Figure 1: Hypothalamus Pituitary- Thyroid axis (Sidhaye, 2012)

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PHYSIOLOGY OF THYROID HORMONE

figure 2: Systhesis of thyroid hormone (Byju, 2022) 10

 THE ROLE OF THYROID HORMONE IN FETAL
AND INFANT DEVELOPMENT

Figure 3: The role of thyroid hormone in fetal and infant development (Klosinska et al., 2022) 11
 ETIOLOGY OF CRETINISM
• Lack of thyroid gland (congenital cretinism )
• Failure of the thyroid gland to produce thyroid hormone (congenital
iodine deficiency syndrome).
• Iodine deficiency in the diet (Endemic cretinism).
• Maternal hypothyroidism.
• Treatment of hyperthyroidism using antithyroid drugs or radioactive
iodine.
(Krishna, 2022)

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PATHOPHYSIOLOGY OF CRETINISM 1/2

figure 4: pathophysiology of cretinism (Devos, 2006),

13
 PATHOPHYSIOLOGY OF CRETINISM 2/2

Figure 5. Fetal thyroid gland maturation. (Klosinska et al., 2022)

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 SIGNS AND SYMPTOMS 1/3
• Puffy face, goiter

• Protruding tongue

• Hoarse cry

• Distended abdomen

• Muscle weakness

• Slow reflexes

• Feeding problem

• Lethargy (Krishna, 2022) 15

 SIGNS AND SYMPTOMS 2/3

• Hypothermia

• Decreased intelligent quotient (IQ)

• Umbilical hernia

• Retardation of sexual attributes in individuals

• Dwarfism or short stature

• Reduced muscle tone and coordination with mild neurological

impairment

• Neonatal jaundice (Krishna, 2022) 16

 SIGNS AND SYMPTOMS 3/3

Figure 6: A baby showing the signs and symptoms of cretinism (Delange, 2005)
17
 COMPLICATIONS

• Abnormal walk

• Muscle spasticity

• mutism

• Autism

• Vision and hearing loss

• Difficulties in memorization and attentiveness

(Delange, 2005)
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THE ROLE OF CLINICAL BIOCHEMISTRY

19
 LABORATORY DIAGNOSIS 1/2
URINARY IODINE TEST

THYROID FUNCTION TEST

• TSH

• T4

• T3

• FREE T3

• FREE T4

• TBG 20
 LABORATORY DIAGNOSIS 2/2
URINARY IODINE METHODS

1. Qualitative method

• Rapid urine iodine test kit

2. Quantitative method

• Urinary iodine micromethod

• Inductively-coupled plasma mass spectrometry (ICP-MS)

21
 URINARY IODINE MICROMETHOD 1/4
SAMPLE COLLECTION:

• The specimen type is urine (24 hours or spot urine).

• Volume required : 1.5-7.0ml.

• Acceptable containers include sterile and prescreened 15ml PP tubes and

5.0ml cryovials.

• Preservatives: 50% acetic acid or refrigerate at 4-8 ºC.

(Mayo clinic, 2023)

22
 URINARY IODINE MICROMETHOD 2/4
PRINCIPLE:

• Urinary iodine Micromethod incorporates two steps of action, i.e. urine digestion
at high temperature and iodine measurement in Sandell-Kolthoff reaction of:

As3+ + I2 As5+ + 2I-

2Ce4+ + 2I- 2Ce 3++ I2

(yellow) (colorless)

(Hassan and Selamat, 2018)

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 URINARY IODINE MICROMETHOD 3/4
Chemicals

• Ammonium persulfate (Digestion)

• Arsenious acid solution (As2O3, NaCl, H2SO4) (Adding As3+ions)

• Ceric ammonium sulfate, H2SO4 Adding (Ce4+ ions)

(Hassan and Selamat, 2018)

24
 URINARY IODINE MICROMETHOD 4/4

25
Figure 7: Procedure for urinary micromethod (Hassan and Selamat, 2018)
 URINARY IODINE TEST REFERNCE RANGE

figure 8: reference range for urinary iodine test 26

(Khazan et al., 2013)
 ICP-MS 1/3

• Inductively coupled plasma mass spectrometry (ICP-MS) is a type of

mass spectrometry that uses an inductively coupled plasma to ionize the
sample. It atomizes the sample and creates atomic and small polyatomic
ions, which are then detected. It is known and used for its ability to detect
metals and several non-metals in liquid samples at very low
concentrations.

(Lee, 2018)

27
 ICP-MS 2/3

Figure 9: ICP-MS Machine (Lee, 2018) 28

 ICP-MS 3/3

Figure 10: ICP-MS procedure (Lee, 2018) 29

 THYROID FUNCTION TEST METHODS

• AUTOLUMO A1000/A1000B ANALYSER

• ELISA

• FLOURCARE

30
 AUTOLUMO A1000/A1000B ANALYSER 1/4

• Autolumo A1000 is a fully automated Chemiluminescence

Immunoassay Analyzer based on CLIA Microparticles platform ,
which is designed for both quantitative and qualitative detection of
biomarkers related to various diseases and dysfunctions.

(Roberts and Sobel,

2013).

31
 AUTOLUMO A1000/A1000B ANALYSER 2/4
PRINCIPLE

Immunoassay analyzer is based on the principle of magnetic particle

chemiluminescence. The materials of patient body fluids and corresponding
reagents are combined with coating magnetic particles to form luminous
compounds. After washing area, the luminous compounds will be separated
into two kinds of substance, one for test, and the other one for waste.

(Roberts and Sobel, 2013).

32
 AUTOLUMO A1000/A1000B ANALYSER 3/4

The chemically emitted light is measured by a selected high sensitive, low

noise photomultiplier which is operating as an ultrafast photon counter. Then
photons produced from the determined under the substrate catalytic will be
counted and the inspection results are qualitative output through the collection
and conversion of photon numbers. Not the number of count, but the relative
light unit are used as unit of the measurement for the raw data.

(Roberts and Sobel, 2013).

33
 AUTOLUMO A1000/A1000B ANALYSER 4/4

Figure 11: An Autolumo A1000 analyser (Roberts and Sobel, 2013).

34
 ELISA 1/4
PRINCIPLE:

In ELISA, various antigen-antibody combinations are used, always

including an enzyme-labeled antigen or antibody, and enzyme activity
is measured colorimetrically.

The enzyme activity is measured using a substrate that changes color

when modified by the enzyme. Light absorption of the product
formed after substrate addition is measured and converted to numeric
values.
(MBL Life Science., 2023)
35
 ELISA 2/4
PROCEDURE

• Coat the micro titer plate wells with antigen.

• Block all unbound sites to prevent false positive results.

• Add sample containing antibody to the wells and incubate the plate at
37°c.

• Wash the plate, so that unbound antibody is removed.

(Klein et al., 2000)

36
 ELISA 3/4

• Add secondary antibody conjugated to an enzyme (e.g. anti- mouse IgG).

• Wash the plate, so that unbound enzyme-linked antibodies are removed.

• Add substrate which is converted by the enzyme to produce a colored

product.

• Reaction of a substrate with the enzyme to produce a colored product.

(Klein et al., 2000)

37
 ELISA 4/4

Figure 12: A microplate showing antigen- antibody reaction (MBL Life Science., 2023)
38
 FLUORECARE 1/3

PRINCIPLE

The fluorecare immunoassay technology allows the quantification of

single or multiple analytes at the same time by measuring the laser-
epifluorescence. The test is performed on a cartridge with specific
parameters.

(sycomed, 2019)

39
 FLUORECARE 2/3

figure 13: A fluorecare cartridge (sycomed, 2019)

40
 FLUORECARE 3/3

Figure 14: Procedure for fluorecare analyzer (sycomed, 2019)

41
 RESULTS
• REFERENCE RANGE
Age TSH T3 T4 Free T3 Free TBG
(mU/L) (ng/dL) (Ug/dl) (pg/mL) T4 (mg/dL)
(ng/dL)

CORD 2.2-10.7 15-75 7.4-13.0 0.2-2.4 0.9-2.2 1.4-9.4

BLOOD
1-4 2.7-26.5 100-740 11.8-22.6 1.8-7.6 2.2-5.3 -
DAYS
4-30 DAYS 1.2-13.1 105-387 9.8-16.6 2.93-5.08 0.9-3.4 1.9-4.5
1-12 1.9-4.4
MONTHS 0.6-7.3 105-245 7.2-16.5 2.67-5.21 0.9-2.3

(Lem et al.,2012)
Table 1: Reference range for thyroid function test
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 OTHER LABORATORY INVESTIGATIONS

• SERUM BILIRUBIN: Increase levels of serum bilirubin

• SERUM CREATINE KINASE: Increased levels associated with

myopathy

• LIPID PROFILE: Hypercholesterolemia and hypertriglyceridemia

• SERUM ELECTROLYTES: Hyponatremia

( Thahir,2023)

43
 CRETINISM TREATMENT

• Once diagnosed, treatment should be started within 1-2 weeks of life.

• Treatment should be started by the onset of symptoms, if

developmental abnormalities and mental retardation start to appear, it
will not be reverse even with thyroid hormone replacement therapy.

• The recommended dose is 10 to 15 μg per kg of body weight. The

dose should be increased as the age progresses.
(Byjus, 2022)
44
 CONCLUSION

Cretinism is a condition of severe physical and mental retardation due to

iodine deficiency, specifically deficiency of thyroid hormones during
early pregnancy. It is typically diagnosed during infancy or childhood.
The congenital hypothyroid child may appear normal at birth or present
with slightly increased head size due to myxedema of the brain.

45
 RECOMMENDATION
• FMC Bida Chemical pathology Unit should advocate for Thyroid function test for newborns
seen with certain signs and symtopms of cretinism such as neonatal jundice and umbilical
hernia etc..

• Mostly importantly, FMC Bida should introduce Urinary Iodine Test as part of Ante-natal
screening for pregnant women.

BECAUSE PREVENTION IS NOT ONLY BETTER BUT CHEAPER

THAN CURE.
46
 REFERENCES
Byju, R. (2022). Cretinism: meaning, causes, symptoms, congenital and Endemic
Cretinism. British Journal of Paediatric. 13(1):1.
Delange, F. (2005). Neonatal thyroid screening as a monitoring tool for the control of
iodine deficiency. Acta Paediatric Supply. 88(432):21-24.
Devos. (2005). How should we be treating children with congenital hypothyroidism?.
Journal of Pediatric Endocrinology and Metabolism. 20(5):559-578.
Hussain, H., Selamat, R. (2018). Understanding Quality Control with Urinary Iodine
Estimation. IntechOpen Reviews. 72-74.
Jibril, M., Abbbiyeseku, F., Aliyu, I., Randawa, A.J., Adamu, R., Akuyam, S., Manu,
M., Suleiman, H.M., Adamu, S., Yusuf, R., Mohammed A. (2016).
Nutritional iodine status of pregnant women in Zaria, North-Western Nigeria. Sub-
Sahara African Journal of Medicine. 1(3):41-44.
Kayode-Adedeji, B., Alikah, S. (2015). Congenital hypothyroidism in a five year old
Nigerian girl: A case report. International Journal of Basic, Applied and
Innovative Research. 4(1):15-19. 47
 REFERENCES
Khazan, M., Azizi, F., Hedayati, M. (2013). A review on Iodine determination methods
in salt and biological samples. Scimetripe Reviews. 1(1):1-4.
Klein, A.H., Meltzer, S., Kenny, F.M. (2000). Improved prognosis in congenital
hypothyroidism treated before age three months. Journal of Pediatry. 81(5):912-915.
Klosinska, M., Kaczynska, A., Ben-Skowronek, I. (2022). Congenital hypothyroidism
in preterm Newborns- The challenges of Diagnostics and treatment. Frontiers in
Endocrinology. 10(13):860-862.
Krishna, V. (2022). Cretinism. Biology of Cretinism.
Lee, H.C. (2018). "Review of inductively coupled plasmas: Nano-applications and
bistable hysteresis physics". Applied Physics Reviews. 5 (1):11-18.
Lucas, P., Davide, C., Daniela, C. (2010). Genetics and phenomics of hypothyroidism
due to TSH resistance. Molecular and Cellular Endocrinology.
Mayo clinic. (2023). Urine Preservatives—Collection and Transportation for 24-Hour
Urine Specimens. https://www.mayoclinic.org/Urine Preservatives—Collection and
Transportation for 24-Hour Urine Specimens. 48
 REFERENCES
Rahul, B., Damor, J.C., Sukesha, G., Jayant, P., Kosambiya, J.K. (2017). Urinary iodine
excretion in urine samples among children in Dang district, Gujarat.
International Journal of Community Medicine and Public Health. 4(1):2339- 2343.
Sidhaye, A.R., Wondisford, F.E. (2012). Handbook of Neuroendocrinology. Academic
Press. 1:685-706.
Sycomed, A. (2019). FLUORECARE quantitative. Sycomed Reviews.
Thahir, R. (2023). Congenital hypothyroidism. The rish academy Review.

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