placenta and amniotic fluid

Placenta and amniotic fluid

Embryogenesis

Fertilization

The Nuclei Fuse Together

What happens now?

Development of the zygote, the study of which is known as embryology or developmental biology.

The zygote undergoes a series of mitotic cell divisions called cleavage.

The stages of development are: Fertilized ovum (zygote) 2-cell stage 4-cell stage 8-cell stage Morula Blastula Early Gastrula Late Gastrula

Cleavage (divide via mitosis) forms the 2 cell stage

They split again to form the 4 cell stage

And again to form the 8 cell stage…

And eventually form a Morula

Next it becomes a blastula

58 cell stage 5 embryo producing

cells+53 cells form trophoblast

107 cell blastocyst (8+99)

no larger Released from zona

pellucida

And next, a gastrula

The Regents Diagram…

1. Sperm and ovum

2. Zygote (fertilized ovum)

3. 2-cell stage

4. 4-cell stage

5. Morula

6. Blastula

7. Gastrula

Differentiation (Organogenesis)

Organogenesis is the formation of the organs (Organo = organs, genesis = creation)

Arises from the layering of cells that occurs during gastrula stage

The layers are germ layers; they have specific fates in the developing embryo:

Differentiation (Organogenesis)

Endoderm The innermost layer Goes on to form the gut

Mesoderm In the middle Goes on to form the muscles,

circulatory system, blood and many different organs

Ectoderm The outermost Goes on to form the skin and

nervous system

Late Gastrula

Ectoderm

Endoderm

Mesoderm

Where does this all take place?

Decidual structure

Decidua is an analogy to deciduous leaves (to indicate that it is shed after childbirth) Decidua basalis Decidua capsularis decidua parietalis

or vera

Fusion of capsularis and parietalis at 14-16wks causes functional obliteration of the uterine cavity

8th week

1. Decidua parietalis2. Decidua capsularis3. Decidua basalis4. Uterine cavity

The decidua consists of various parts, depending on its relationship with the embryo: Decidua basalis, where the

implantation takes place and the basal plate is formed. This can be subdivided into a zona compacta and a zona spongiosa (where the detachment of the placenta takes place following birth).

Decidua capsularis, lies like a capsule around the chorion

Decidua parietalis, on the opposite uterus wall

5. Smooth chorion (laeve)

6. Chorionic villi7. Amniotic cavity8. Decidua capsularis

and parietalis, grown

together

Decidual reaction

polygonal or round Round and

vesicular nucleus Cytoplasm clear,

basophilic Pericellular

membraneWalls around themselves and around the fetus

Decidual blood supply

Spiral arteries in the parietalis retain smooth muscle wall and epithelium

Cytotrophoblast invasion of spiral arteries and arterioles -vessel wall in the basalis destroyed-not responsive to vasoconstrictors

Decidual histology

True decidual cells Maternal bone marrow derived cells Decidual NK cells

Secrete cytokines Express angiogenic factors

Basalis-mainly arteries and widely dilated veins(glands virtually disapperared)

Invasion by trophoblasts

Decidual prolactin

Paracrine between maternal and fetal tissues Amnionic 10,000ng/ml Maternal 150-200ng/ml Fetal 350ng/ml

Role ? Transmembrane solute and water

transport Stimulate T-cells Regulates angiogenesis

Implantation

The embryo implants in the wall of the uterus on about the 7th day of development

Blastocyst implantation

Apposition- days 20-24 of cycleendometrium primed

by E&P Adhesion modification in expression

of cellular adhesion molecules (integrins)

Invasion

12-day Human Embryo

Gas and nutrient exchange system Embryo is nourished in the first weeks

through simple diffusion Utero-placental circulation

system in which the circulation systems of the mother and of the embryo get closer together, thus allowing an exchange of gases and metabolites via diffusion.

Maternal and fetal blood never come into direct contact with each other.

Trophoblast differentiation cytotrophoblast

inner layerwell

demarcated Syncytiotrophoblast

outer layermultinucleated

After implantation

Villous trophoblast• chorionic villi

Extra-villous trophoblast

• interstitial• endovascular

Lacunar stage

Through the lytic activity of the syncytiotrophoblast the maternal capillaries are eroded and anastomose with the trophoblast lacunae, forming the sinusoids.

Lacunae communicate with each other and form a single, connected system that is delimited by the syncytiotrophoblast and is termed the intervillous space.

Lacunar stage

primary villi

D11-13

Syncytiotrophoblast penetrated by cords of cytotrophoblast

primary villi

Secondary villi

After the 16th day The extra-embryonic mesoblast also

grows into this primary trophoblast villus, which is now called a secondary villus and expands into the lacunae that are filled with maternal blood.

The ST forms the outermost layer of every villus.

Secondary villi

Tertiary villi

At the end of the 3rd week the villus mesoblast differentiates into connective tissue and blood vessels.

Villi that contain differentiated blood vessels are called tertiary villi

The EEM remains in this stage, still surrounded by cytotrophoblast. The outer envelope of the villus is still formed by the ST

Tertiary villi

Free villi

After 4th month the cytotrophoblast in the tertiary villi disappear slowly

the villi divide further and become very thin.

1. Anchoring villus2. Septum3. Syncytiotrophoblast (ST)4. Cytotrophoblast (CT)5. Remainder of the cytotrophoblast

layer 6. CT in the spiral artery wall

A-Basal plate and myometrium

B-Chorionic plate

Ageing of the placenta

A. Langhans' fibrinoid layer

B. Rohr's fibrinoid layer

C. Nitabuch's fibrinoid layer

Cytotrophoblastic invasion

destruction of the smooth muscle layer

partial replacement of the endothelial cells

change in elasticity of the spiral arteries,

Absent in preecclampsia and intra-uterine growth retardation.

excessive proliferation of the cytotrophoblast can lead to tumor formation, especially to a chorion carcinoma.

Development of the Placenta

Placental tissue structureChorionic plate Basal plate

1 Amnion2 Extra-embryonic mesoblast3 Cytotrophoblast4 Syncytiotrophoblast

5 Zona compacta6 Zona spongiosa7 Decidua basalis8 Myometrium

Fetal circulation system

1. Umbilical arteries

2. Umbilical vein

3. Fetal capillaries

A network of fetal capillaries (2 to 8) is found in each villus;

20 to 40 first order stem villi exist from each one of which 20 to 50 second and third order daughter villi arise.

Maternal circulation system

1. Spiral arteries

2. Uterine veins

3. Intervillous spaces

Spiral arteries (branches of the uterine arteries)

High pressure At the level of the

placenta (intervillous spaces), therefore, maternal blood is to be found at times outside the vessel network.

Blood pressure values and oxygen distribution in the intervillous spaces

Maternal blood is pumped with high pressure and leaves via the uterine veins. At the level of the placenta (intervillous spaces), therefore, maternal blood is to be found at times outside the vessel network.

Development of the placenta (> 4th month)

1. Decidual tissue2. Syncytiotrophoblast3. Cytotrophoblast islands4. Septum

The cytotrophoblast islands move into the periphery of the cotyledons and, together with the decidual tissue, are involved with formation of the placental inter-cotyledon septa.

Fibrinoid degradation

The villus stems of the placenta lengthen considerably towards the end of the pregnancy and the fibrinoid deposits (extra-cellular substance made up of fibrin, placental secretions and dead trophoblast cells), accumulate in the placenta

Fibrinoid degradation

structurally and chemically closely related to fibrincan take up a maximum of 30% of the placental volume without affecting its function. When these deposits are massive and block one or more vessels to the villi, they form white infarcts,Functional importance

sealing effects, Immunologic "barrier“ anchoring of the

placenta.

A. Subchorial Langhans' layerB. Rohr's layerC. Nitabuch's layer

Placental barrier

First trimester

1. Intervillous space2. Syncytiotrophoblas

t3. Cytotrophoblast4. Villus mesenchyma5. Fetal capillaries6. Hofbauer

macrophages

Placental barrier

2nd trimester

1. Intervillous space2. Syncytiotrophoblast3. Cytotrophoblast4. Villus mesenchyma5. Fetal capillaries6. Hofbauer

macrophages

Placental barrier

3rd trimester

1. Intervillous space

2. Placental barrier of a terminal villus

3. Fetal capillaries

4. Merged basal membranes

5. Endothelial cells

6. Rare cytotrophoblast cells

7. Basal membrane of the capillaries

8. Basal membrane of the trophoblast portion

9. Syncytiotrophoblast with proliferation knots (nuclei rich region)

Placenta

Placental transport

Passive transport Simple diffusion:

non-polar molecules fat-dissolvable substances (e.g., diffusion of

oxygen, carbon dioxide, fats and alcohol). Water enters the placenta through specialized pores (see osmosis).

Osmosis: theaquaporines or water channels, proteins localized within the plasma membrane.

Simplified transport: transition from the side with higher concentration to the one with lower concentration with the help of transport molecules (e.g., glucose).

Placental transport

Active transport: Transport through the cellular membrane against a concentration gradient using energy (Na+/K+ or Ca++)

Vesicular transport (Endocytosis / Exocytosis): Macro-molecules are captured by microvilli and absorbed in the cells or repelled (immunoglobulin).

averages 22 cm (9 inch) in length 2–2.5 cm (0.8–1 inch) in thickness weighs approximately 500 grams dark reddish-blue or crimson color Umbilical cord of approximately 55–60 cm,

which contains two umbilical As and one umbilical V and has an eccentric attachment.

On the maternal side, these villous tree structures are grouped into lobules called cotyledons

Placental functions

The placental exchange surface is enlarged from 5 m2 at 28 weeks to roughly 12 m2shortly before delivery!

Placental functions

Breathing function Nutritive and excretory functions Placenta and the immunological

barrier Protein transfer Protective function Endocrinal function

Breathing function

The placenta, which plays the role of "fetal lungs", is 15 times less efficient (with equivalent weight of tissue) than the real lungs.

The supply of the fetus with oxygen is facilitated by three factors: difference of oxygen concentration and partial

pressure within the feto-maternal circulation system

higher affinity of fetal hemoglobin (HbF) for oxygen

Bohr effect

Nutritive and excretory functions

Water diffuses into the placenta along an osmolar gradient. The water exchange increases during the pregnancy up to the 35th week (3.5 liter / day).

The electrolytes follow the water, whereby iron and calcium only go from mother to child.

Glucose is the fetus' main source of energy and passes the placenta via simplified transport. At the level of the trophoblast the placenta can synthesize and store glycogen in order to satisfy local glucose requirements through glycogenolysis.


Peptides and amino acids via active transport and thus insure the fetus' own protein synthesis.

Amino acids, precursors of fetal protein synthesis, stem from the metabolism of the maternal proteins. The placental transport is facilitated by the influence of hormones, e.g., GH (growth hormone) and TSH (thyroid stimulating hormone) against a concentration gradient (2-3 times higher in the fetus as in the mother).

Lipids and triglycerides are decomposed in the placenta, where new lipid molecules are synthesized.

Cholesterol passes through the placental membrane easily, just like its derivates: e.g., steroid hormones.


Water-soluble vitamins easily pass through the placental membrane. The amount of the fat soluble vitamins (A,D,E and K) in the fetal circulation is, on the other hand, quite low. Vitamin K plays an important role in blood coagulation and is applied to the child immediately after birth, in order to prevent hemorrhages.

Placental exchange processes are also involved in the removal of waste products from the fetal metabolism. They cross over into the maternal blood in order to be excreted by the mother (urea, creatinine, ureic acid).

Placenta and the immunological barrier The fetus is not rejected even though its set of

chromosomes differs from that of its mother and halfway represents an allogenic transplantation

Fetal tissue and especially that of the placenta that stand in direct contact to the maternal organismproduce no tissue antigens

HLA -G antigens, which do not distinguish between individuals, occurs through the extravillous cytothrophoblast. The HLA-G antigen takes over anti-viral and immunosuppressive functions as well as non-immunologic tasks.

Placenta and the immunological barrier

In addition, the placenta blocks cytotoxic maternal cell effects by secreting various factors. The insufficiency of these mechanisms may be responsible for immune-dependent miscarriages.

some steroid hormones (e.g., progesterone) have an immunosuppressive effect on the lymphocytes of the pregnant woman. Progesterone (the concentration of which is especially elevated during pregnancy) seems to play an important immunosuppressive role that is mediated by the PBIF protein (Progesterone Induced Blocking Factor).

Protein transfer

The maternal proteins do not traverse the placental barrier, with the exception of immunoglobulin (IgG). Through pinocytosis of syncitiothrophoblast cells the mother thus transfers to the fetus the variety of IgG that she has synthesized during her life. This transfer occurs mainly towards the end of pregnancy. Thereby the fetus obtains a passive immunity that protects it against various infectious diseases in the first six months of its life. The other immunglobulins, mainly IgM proteins, do not pass through the placental barrier.

Protein transfer

Other proteins:Transferrin is another important maternal protein that, as the name indicates, transports iron. On the surface of the placenta specific receptors exist for this protein, which, by means of active transport, enters into fetal tissue.

Protein can also be transferred from the fetus to the mother; alpha-fetoprotein (the concentration of which is elevated in several fetal abnormalities) can be detected in the maternal circulation system.

Maternal or placental polypeptide hormones do not enter the fetal circulation system.

Protective function

Sexually transmitted diseases: After the 5th month of pregnancy treponema

pallidum bacteria, the syphilis pathogen, can pass through the placental barrier.

HIV transmission from the mother to the fetus amounts to roughly 15 to 25%. It depends on the viremia status of the mother. Anti-HIV treatment during the pregnancy and birth as well as

further treatment of the newborn during the first few weeks. Birth via caesarian section No breastfeeding of the child

When all of these measures have been carried out the risk of infection for the baby can be reduced to below 1%.

Protective function

Fetotoxic infections: The rubella virus may be responsible for a

miscarriage during pregnancy (before the first month), for embryopathies (when the virus invades between the 1rst and 3rd month) or for fetopathies (after the 3rd month).

Toxoplasmosis is harmless for the mother, but can cause severe anomalies in the fetus.

Listeriosis can be responsible for miscarriages, intrauterine death or neonatal sepsis due to transplacental infection or for secondary late meningitis due to a contaminated birth passage.

Protective function

Fetotoxic infections: The cytomegalovirus is generally the cause

of infections that remain subclinical. It can also be responsible for miscarriages as well as for microcephaly and growth retardation. The infection happens transplacental or during birth.

The parvovirus B19 is responsible for aplastic crises in utero (marked decrease of blood cells).

Protective function

In addition, the placenta also presents an incomplete barrier against certain injurious effects of drugs: Antibiotics and corticoids can pass through the placental barrier. Depending on their size, certain steroid hormones get through as well.

Endocrinal function

The placenta and especially the syncytiotrophoblast can be seen as a large endocrine gland.

Before implantation hormone production is ensured through ovarian and hypophysial hormones.

At the beginning of the pregnancy the synthesis of estrogen and progesterone is ensured by the corpus luteum graviditatis that is maintained by the human chorion-gonadotropin (HCG), a product of the trophoblast. The activity of the corpus luteum decreases progressively with the beginning of the 8th week in order to be entirely replaced by the placenta at the end of the 1st trimester

During the pregnancy the hormone concentration in the maternal blood is regulated by the cooperation of the placental, hypophysial and fetal suprarenal hormones as well as hormones from the gonads

Endocrinal function

a Placenta

b Fetal suprarenal glands

c corpus luteum graviditatis

Implantation and form anomalies placenta previa

Implantation and form anomalies Ectopic, i.e., extra-uterine

pregnancies

Implantation and form anomalies

Implantation and form anomalies Velamentous insertion

Implantation and form anomalies marginal

Implantation and form anomalies Eccentric insertion

Implantation and form anomalies• Placenta multilobata

CLINICAL BIOCHEMISTRY AMNIOTIC FLUID


Implantation and form anomalies placenta succenturia

Implantation and form anomalies bilobate when both segments of the placenta

are almost equal in size (right on the figure) and succenturiate when there is a greater difference (left on the figure). When there is not such a connection, the placenta is called placenta spuria.

Implantation and form anomalies•Circumvalate placenta


Implantation and form anomalies fenestrated placenta


Implantation and form anomalies Membranaceous placenta

Implantation and form anomalies Disc shaped

placenta

Toxemia of pregnancy

Fetal erythroblastosis

anemia (due to the hemolysis)

splenomegaly (location of the macrophages that destroy the erythrocytes)

hepatomegaly (intensive hematopoesis in order to compensate the hemolysis)

icterus (transformation of hemoglobin of the destroyed erythrocytes into bilirubin)

Inflammation of the placenta Bacterial infections can also strike the

placenta (placentitis) or the fetal membranes (chorioamnionitis). Normally, these infections are transmitted vaginally in the case of an early rupture of the amnion. An infection rarely occurs via the blood, i.e., when the fetal membrane is still intact. Syphilis was earlier a frequent cause for placentitis, also for placental tuberculosis, whereby here the placenta was infected via the blood

Hydatid moleThe hydatid mole pregnancy corresponds to a cystic chorion villus degeneration

Macroscopically, the mole looks like a heap of transparent bubbles, held together by filaments, and supported by a central core.

Hydatid mole

Microscopically, the villus degeneration exhibits no vascularization, a proliferation of trophoblasts (from cytotrophoblasts – Langhans' cells and from syncytiotrophoblasts) and dystrophic alterations of the connective tissue with stroma edema.

The chorion and amnion enclose the embryo

The chorion surrounds the entire embryoThe amnion encloses the embryo and forms an open volume between the embryo & the amnion called the amniotic cavityAmnion provides almost all tensile strength

AMNIOTIC FLUID


Development

Amniogenic cells line the inner surface of trophoblast

Derived from fetal ectoderm of the embryonic disc

Amnion & Amniotic Fluid

Composition of Amniotic Fluid 99% H2O Un-disolved material

Organic & inorganic salts Pregnancy advancement changes its

composition Meconium & urine

Amniotic Fluid

Before 20 weeks gestation – AF is an ultrafiltrate of maternal

serum Maternal & AF osmolality, sodium,

urea, and creatinine are roughly equal.

At term Volume = 900cc Reflective of fetal renal function. Progressively hypotonic. Contains fetal debris: squamous

cells, mucin, lanugo.

Amniotic Fluid

Amniotic fluid surrounds the fetus during intrauterine development.

This fluid cushions the fetus against trauma,

Has antibacterial properties to lessen infections,

Reservoir that may provide a short-term source of fluid and nutrients to the fetus.

Amniotic Fluid

Amniotic fluid are required for the fetal musculoskeletal system to develop normally, for gastrointestinal system development, and for the fetal lungs to develop.

It is not surprising to find that oligohydramnios and polyhydramnios are associated with increased rates of perinatal morbidity and mortality.

Sources of amniotic fluid

The two primary sources of amniotic fluid are fetal urine and lung liquid, with an additional small contribution due to secretions from the fetal oral-nasal cavities.

Fetal urine is a major source of amniotic fluid in the second half of pregnancy.


Urine production Approximately 110/ml/kg every 24 hours at 25 weeks to approximately 190 ml/kg every 24 hours at 39 weeks

At term, the current best estimate of fetal urine flow rate may average 700-900 ml/day.


The fetal lungs are the second major source of amniotic fluid during the second half of gestation.

Studies in near-term fetal sheep have shown that there is an outflow from the lungs of 200-400 ml/day


The inward transfer of solute across the amnion with water following passively is the most likely source of amniotic fluid very early in gestation

Part of AFV may be derived from water transport across the highly permeable skin of the fetus during the first half of gestation, at least until keratinization of the skin occurs around 22-25 weeks.

Routes of amniotic fluid removal

The two primary routes of amniotic fluid removal are fetal swallowing and absorption into fetal blood perfusing the fetal surface of the placenta.

Fetal swallowing plays an important role in determining AFV during the last half of gestation.

Routes of amniotic fluid removal

The fetus begins swallowing at the same gestational age when urine first enters the amniotic space, that is around 8-11 weeks.

It is estimated that the volume of amniotic fluid swallowed in late gestation averages 210-760 ml/day

Intermembranous & transmembranous pathways

As a further pathway, rapid movements of both water and solute occur between amniotic fluid and fetal blood within the placenta and membranes; this is referred to as the intramembranous pathway.

Movement of water and solute between amniotic fluid and maternal blood within the wall of the uterus is an exchange through the transmembranous pathway

Amniotic fluid volume

Amniotic fluid volume

The rate of change in AFV is a strong function of gestational age.

There is a progressive AFV increase from 30 ml at 10 weeks’ gestation to 190 ml at 16 weeks and to a mean of 780 ml at 32-35 weeks, after which a decrease occurs

The decrease in post-term pregnancies has been found to be as high as 150 ml/week from 38 to 43 weeks

Individual amniotic fluid volumes from acollection of 705 measurements in patients with anormal pregnancy outcome

Regulatory mechanisms act at three levels:

Placental control of water and solute transfer.

Regulation of inflows and outflows from the fetus: fetal urine flow and composition are modulated by vasopressin, aldosterone, and angiotensin II in much the same way as they in adults.

Maternal effect on fetal fluid balance: during pregnancy, there is a strong relationship between maternal plasma volume and AFV,

Measurement of amniotic fluid volume

Single vertical pocket

Amniotic fluid index

Oligohydramnios

Diminished amniotic fluid volume (AFV)

Amniotic fluid volume of less than 500 mL at 32-36 weeks' gestation - Amniotic fluid volume depends on the gestational age; therefore, the best definition may be AFI less than the fifth percentile.

Single deepest pocket (SDP) of less than 2 cm

Amniotic fluid index (AFI) of less than 5 cm or less than the fifth percentile

Oligohydramnios-causes

Fetal Chromosomal

anomalies Congenital

abnormalities Growth restriction Demise Post-term pregnancy Ruptured membranes

Placental Abruption TTTS

Maternal Uteroplacental

insufficiency Hypertension Pre-ecclampsia Diabetes

Iatrogenic PG synthesis inhibitors ACE inhibitors

Idiopathic

Congenital anomalies associated with oligohydramnios Amnionic band syndrome Cardiac

Fallots tetralogy Septal defects

CNS Holoprosencephaly Meningocele Encephalocele microcephaly

Cloacal dysgenesis Chromosomal

Triploidy Trisomy 18 Turner syndrome

Cystic hygroma Diaphragmatic hernia Genitourinary

Renal dysgenesis/aplasia Urethral obstruction Bladder exystrophy Meckel gruber syndrome Uretro-pelvic junction obstruction Prune belly syndrome

Hypothyroidism Skeletal TRAP sequence TTTS VACTERL association

Oligohydramnios

Fetal mortality rates as high as 80-90% have been reported with oligohydramnios diagnosed in the second trimester.

Midtrimester PROM often leads to pulmonary hypoplasia, fetal compression syndrome, and amniotic band syndrome.

Oligohydramnios is a frequent finding in pregnancies involving IUGR and is most likely secondary to decreased fetal blood volume, renal blood flow, and, subsequently, fetal urine output.

AFV is an important predictor of fetal well-being in pregnancies beyond 40 weeks' gestation

AFV is a predictor of the fetal tolerance of labor,

Oligohydramnios

Ultrasonography diagnosis is confirmed ultrasonography of the

fetal anatomy

Sterile speculum examination Pooling in posterior

fornix Nitrazine paper turns

blue arborization or ferning

pattern

amnioinfusion

Polyhydramnios

Polyhydramnios is the presence of excess amniotic fluid in the uterus.

Deepest vertical pool is more than 8 cm

AFI is more than 95th percentile for the corresponding gestational age.

The incidence is 1-3% of all pregnancies.

About 20% are associated with fetal anomalies.

The diagnostic approach to polyhydramnios consists of

(1) physical examination of the mother with an investigation for diabetes mellitus, diabetes insipidus, and Rh isoimmunization; (2) sonographic confirmation of polyhydramnios and assessment of the fetus; (3) fetal karyotyping; and (4) maternal serologic testing for syphilis.

polyhydramnios

Maternal hyperglycemia GIT

anomalies(obstructive) Esophageal atresia Tracheoesophageal fistula Duodenal atresia

Nonimmune hydrops CNS anomalies

Anencephaly Open spina bifida

Thoracic malformations Diaphragmatic hernia

Congenital infections Syphilis, hepatitis

Chromosomal anomalies High output Cardiac

failure Fetal anemia Sacrococcygeal teratoma chorioangioma

Fetal polyuria Fetal

pseudohyperaldosteronism

Fetal bartter Nephrogenic diabetes

insipidus Placental chorioangioma Maternal substance

abuse

Amniotic fluid testing

Chromosome and DNA analysisBiochemistry Fetal infectionsRh disease and other

alloimmunisationLung maturityChorioamnionitisObstetric cholestasisFetal therapy- decompression

severe oligohydramniosmultifetal pregnancy

reductionthroxine therapy

Prenatal diagnosis

Alpha fetoprotein

Measurement of AFP in maternal serum and amniotic fluid is used extensively for the prenatal detection of some serious fetal anomalies.

AFP Biochemistry

AFP is produced initially by the fetal yolk sac in small quantities and then in larger quantities by fetal liver as the yolk sac degenerates.

Trace amounts are also produced in the fetal gut and kidneys.

AFP Biochemistry

Concentrations of AFP in fetal serum Early in embryonic life:1/10

the concentration of albumin in fetal serum

16 weeks gestation:3,000,000 ng/ml

At term:declines steadily to 5000 to 120,000 ng/ml

AFP Biochemistry

The rise and fall in concentration of AFP in the amniotic fluid roughly parallels that in the fetal serum but lower in concentration 20,000 ng/ml at 16 weeks

gestation

Clinical significance of AFP

Maternal serum and amniotic fluid AFP are useful tests for detecting some serious fetal anomalies

Maternal serum AFP is elevated in 85% to 95% of cases of fetal open neural tube defect and is low in about 30% of cases of fetal Down’s syndrome.

Acetyl cholinesterase

A useful adjunct in the diagnosis of neural tube defects is the measurment of acetylcholinesterase (AChE,EC 3.1.1.7) in amniotic fluid

The usual technique for identification of AChE is polyacrylamide gel electrophoresis.

Acetyl cholinesterase test sensitivity

A study of more than 5000 patients reported that determination of AChE by electrophoresis had specificity of 99.76% and following sensitivities:

Anencephaly,97%

Open spina bifida,99%

Abdominal wall defects,94%


Testing amniotic fluid for AFP and AChE can predict open neural tube defects more accurately than maternal serum screening.

Patient with unexplained high maternal serum AFP levels and normal ultrasonography findings should be offered amniotic fluid testing.

Any patient who has had a child with a neural tube defect has 3% to5% risk for recurrence and also should be offered amniotic fluid AFP testing

Any elevation of AFP in amniotic fluid should lead to AChE analysis


Testing should be performed at or before 16 weeks gestation.

Determination of fetal karyotype is also reasonable.

AF and Respiratory distress syndrome (RDS)


Respiratory distress syndrome (RDS) was associated with a significant mortality rate approaching approximately 30%.

In the 1950s, it was discovered that the resistance of pulmonary alveoli to collapse during expiration was mainly caused by the presence of a surface tension-lowering material lining the alveolus (surfactant).

As the lungs develop, significant quantities of surfactant are washed out of the fetal lung and accumulate in the amniotic fluid.


all of the available biochemical tests for fetal lung maturity rely on the amniotic fluid content of surfactant

adult mature surfactant is approximately 80% phospholipids, about 10% protein, and about 10% neutral lipids (primarily cholesterol).

The major species of phospholipid in surfactant is phosphatidylcholine (also referred to as lecithin), which accounts for 80% of the total phospholipid

Surfactant lipid Composition

L/S ratio test

The L/S ratio test remains one of the most commonly used tests, and one of the standardized tests against which all other tests are compared.

With a L/S ratio of 1.5-1.9, approximately 50% of infants will develop RDS. Below a ratio of 1.5, the risk of subsequent RDS increases to 73%.

One of the major disadvantages of the L/S ratio is the inability to use this test in the setting of contaminated amniotic fluid. Both blood and meconium staining of amniotic fluid have been found to interfere with L/S ratio determinations.

PG determinations:

It is found that the false-positive rate for PG determination was 1.8%. This rate is significantly lower than the false-positive rate they found for the L/S ratio(5%)

PG performs much better than the L/S ratio in predicting babies who will develop RDS. Finally, PG determinations accurately predict pulmonary maturity and give a better indication of pulmonary immaturity than does the L/S ratio

Saturated Phosphatidylcholine

Saturated Phosphatidylcholine has been found to predict pulmonary maturity

Respiratory distress syndrome was correctly predicted 55.5% of the time by L/S ratio and 82% of the time by SPC.

Pulmonary immaturity = an SPC <500 μg/dl

In addition, the SPC was found to be valid in the presence of blood and meconium, whereas the L/S ratio was not.

The lung profile includes the L/S ratio, desaturated lecithin, PG and PI concentrations.

lung profile help to form a clearer picture of fetal lung development

The L/S ratio had a false-positive rate of 3%-5%, which was reduced to less than 1% with the combined lung profile test

Lung Profile

Microviscosimeter

Microviscosimeter testing measures surfactant associated with a phospholipid membrane using fluorescent dye techniques.

The microviscosimeter commonly used in the fetal lung maturity analyzer or FELMA machine.

Surfactant/Albumin Ratio

A recently introduced TDx FLM assay is an automated fetal lung maturity test based on the principle of fluorescent polarization used previously with the microviscosimeter.

A surfactant albumin ratio of 50-70 mg surfactant/g of albumin has been considered mature in most studies

The TDx test correlates well with the L/S ratio and has few false-immature results, making it an excellent screening test

It only requires approximately 1 ml of amniotic fluid and the test can be performed in less than an hour,

Shake test

this test use the principle that when ethanol is added to amniotic fluid, the nonsurfactant foam causing substances in amniotic fluid are removed.

any stable foam layer that persists after shaking is due to the presence of surfactant in a critical concentration.

when serial dilutions of ethanol are used, the surfactant can be quantified.

it is found that the shake test was comparable to the L/S ratio and had a high predictive value for RDS when applied to uncontaminated amniotic fluid.

Tap Test

the tap test examines the ability of surfactant within amniotic fluid to break down bubbles within an ether layer.

the test is performed on 1 ml of amniotic fluid mixed with a drop of 6N hydrochloric acid and 1.5 ml of diethylether

the tube is tapped 4 times and examined for the presence of bubbles within the ether layer.

in mature samples, the bubbles quickly breakdown, whereas in immature amniotic fluid specimens more than 5 bubbles persist in the ether layer.

this rapid test was comparable with the phospholipid profile

Visual Inspection

The basis is whether or not newspaper could be read through the amniotic fluid sample, that is, was the fluid too turbid to read text through.

with clear fluid (readable newsprint) the sensitivity of an immature result is 98%.

Optical Density at 650 nm

with a OD 650 value of 0.15 or greater, the L/S ratio was always greater than 2.0 when the OD 650 was less than 0.15, only 6% of L/S ratios were greater than 2

Diabetes and pulmonary maturity

Amniotic fluid and renal maturity

AF assesment and Renal maturity

The fetal kidneys start to develop during the 4th and 5th weeks of gestation and begin to excrete urine into the amniotic fluid at the 8th to 11th week

At the 20th week the fetal kidneys produce most of the amniotic fluid

Renal maturity is defined by the increase in glomerular filtration and by the maturity of renal tubular cells that begin to express various tubular transporters over the months of gestation


Glomerular filtration in the fetal kidney can be assessed by the concentrations of creatinine and urea in the amniotic fluid

Creatinine concentrations of 2 mg/dl represent an age of at least 37 weeks of gestation

The function of the renal tubule system, specifically proximal tubules, can also be assessed by the concentrations of ß2-microglobulin and NAG in the third trimester of gestation


ß2-Microglobulin produced by the fetus is filtered and reabsorbed by proximal tubules, with an expected reduction in its concentrations at week 36 in normal pregnancies.

This reduction can be considered as an index of renal tubular maturation


Analysis of creatinine and urea in amniotic fluid permits an evaluation of renal maturation.

Creatinine values in the amniotic fluid that best represent fetal maturity are 1.5 to 2.0 mg/dl

AF and Bone Healing

AF and Bone Healing

Hyaluronic acid (HA) is a linear polysaccharide with a high molecular weight.

It is found in all extracellular matrices and has the same structure in all species.

If HA is administered during surgery, scar formation is prevented.

HA is known to reduce scar formation by inhibiting lymphocyte migration, proliferation and chemotaxis, granulocyte phagocytosis , degranulation, and macrophage motility

AF and Bone Healing

HA influences and enhances tissue regeneration through its ability to retain large amounts of water.

HA has been reported to increase osteoblastic bone formation in vitro through increased mesenchymal cell differentiation and migration.

AF and Bone Healing

Human amniotic fluid (HAF), obtained by amniocentesis during the second trimester of gestation, contains high molecular weight HA in high concentrations.

It has been showed that HASA (HA-stimulating activator) which is present in HAF, stimulates the wound to increase the production of endogenous HA.

HAF may increase both endogenous and exogenous HA in the application region.

HAF has been reported to enhance new cartilage formation.


Risk and complications

Pain Leakage Hemorrhage Abortion Fetal injury Orthopaedic abnormalities alloimmunisation

placenta and amniotic fluid

Technology