Download 3Dreconstruction

Three-dimensional reconstruction and morphological
quantization of human embryo hearts for virtual
model validation.
J-M. Schleich*, J-L. Dillenseger**, L. Loeuillet***,
J-P. Moulinoux***, C. Almange*.
*
Département de Cardiologie et Maladies Vasculaires.
Hôpital de Pontchaillou. CHR Rennes 35033. France
**
Laboratoire du Traitement du Signal et de l’Image.
Université de Rennes 1. France
***
Laboratoire d’Histologie et d’Embryologie. Faculté de
Médecine, Rennes. France
Introduction (1)
• Most common congenital anomaly observed in lives-borns
• Incidence of congenital heart defects: 5-8 / 1000 newborns
• Abortus and stillbirths: incidence is 5 times higher than in
liveborn infants
• in the last decade, improvements of prenatal diagnosis methods
=> regular decreasing gestational age of abortion
• major malformations / chromosomic anomalies
• Abortion could be realized from 9-10 weeks
(cranio-caudal length : 30 mm)
Introduction (2)
• Before 13 WG, accurate in vivo diagnosis cannot currently
be made in all fetuses with the imaging instrumentation
available.
• Ex vivo or in vitro imaging techniques have not been
adapted to the dimensions of the organs under examination
Introduction (3)
• Before 13 WG, accurate in vivo diagnosis cannot currently
be made in all fetuses with the imaging instrumentation
available.
• Ex vivo or in vitro imaging techniques have not been
adapted to the dimensions of the organs under examination
Introduction (4)
: imaging of the surgically exposed uterus
40-50 MHz / resolution of 50-100 µm / mouse embryos penetration of US between 6-10 mm
Introduction (5)
Confocal microscopy
• Problems of penetration
• especially used for very small specimens
• limited to image only fluorescent signals
Introduction (6) :
µIRM
Introduction (7)
• Complexity of cardiac anatomy and wide spectrum defects of the heart
=> the anatomo-pathological examination remains the gold standard
allowing the making of accurate diagnoses, as well as appropriate
genetic counseling, and the conduct of epidemiological studies.
However, this method reaches its own limits between 9 and 13 weeks
of gestation, when the embryo is between 25 and 70 mm in length. At
this point, the examination of isolated organs is a challenge,
particularly of the heart, which measures between 3 and 8 mm.
Correlation of prenatal US diagnosis and pathologic finding in fetal anomalies
• Comparison between autopsy studies and prenatal diagnosis
(average gestational age: 23 weeks)
– Pediatr Pathology 1989; 9: 1-9
– Pediatr Develop Pathol 1999; 2: 131-42
– Ultrasound Obstet Gynecol 1999; 13: 117-26
– Méd Fœtale Echogr Gynecol 2000; 41: 18-21
– Ann Pathol 2000; 20: 549-57
• main prenatal diagnosis is « modified » in 20-46% of cases
(especially with polymalformatic syndromes)
• genetic counselling / diagnosis of a syndrome /
determination of etiology or pathogenetic mechanism /
interpretation of severity of the anomaly
Introduction (8)
The aim of this report is to present a methodology which can be applied routinely,
using standard histologic section, enabling the reconstruction, visual estimate
and quantitative analysis of the human embryonic cardiac structures.
This is the first description of 3-dimensional (3-D) reconstruction allowing a
diagnostic analysis of fetal human hearts ≤ 13 weeks of age
Method (1)
• Study approved by bioethic medical committee
• abortion specimens fixed in 4% paraformaldehyde
• visual examination of embryo and length
measurements
• « limited » autopsy is performed
• internal examination requires the dissecting
microscope with camera
• straight midline incision / chest wall removed
• visceral situs noted : thoracic organs removed together
• heart is isolated / weight - size are mesured
Method (2)
The heart specimen used in our study
• human abortus fetus of 68 mm crown/rump length
• 11 WOA (« 13 SA ») / spontaneous miscarriage
• normal development / normal external appearance
without any malformation
• heart weight = 0.2 gr / sized = 9 X 8 mm
Method (3)
• Deshydrated with ethanol
• embedded in paraffin
• serial 10 µ-thick transversal section (Leica RM
2145 microtome) (5300 slices)
• 1 / 10 was stained with haematoxylin-eosin
• the colored slices digitalized on optical microscope
(IKAROS 3 V 4.33 / Metasystems)
Method (4): inversion-cleaning-labeling
OriginalCleaned
picture picture
Labeling picture
Inverted scale
Method (5) : calibration
digitalized scale
pixel = 22, 3 µm
Method (6) : half-automatic matching technics
1/ Matching method based on the geometrical moments
2/ Interactive own-developed fitting package
Method (7) : half-automatic matching technics
1/ Matching method based on the geometrical moments
2/ Interactive own-developed fitting package
Result (1)
Result (2) :
Volume formation / interpolation
Stacking the sections / resampling / interpolation of grey scale slice
Result (3) : Volume formation / visualisation
Result (4) : 2D Visualisation
frontal
sagittal
horizontal
Result (5) :
Volume formation
Result (6) : 3D Visualisation
Interactive manipulation of the heart
(VRML format )
Result (7) : 3D Quantification
Internal volumes
/ voxel = 22, 3 µm
– Ventricles : 28.5 mm3
– Atria : 40.2 mm3
Conclusion (1)
•
Transmission of the digitalized data
•
Visual perception improved by addition of 3rd dimension
•
New approach of the morphological analysis
– Cardiac volume enterely reconstruted
– Exploration of the volume even by transparency
– 2D and 3D quantifications
– Creation of sections in any plane : The hope (the dream) of being able to section
the specimen as many times as needed in all spatial dimensions, and to finally
reexamine it in its entirety has finally be fulfilled.
•
Experimental modelisation
Conclusion (2) : Improvements
Episcopic Fluorescence Imaging Capturing
–
–
–
–
–
–
exact 3D reconstruction from serial sections
« on block » staining procedure
picture ’s acquisition before section
allows automatic contour finding
no disorientation and no distorsion of the sections
but
• necessity of specific stainings
• limited final resolution
• variability of staining ‘ intensity
6- WOA embryo (lenght : 13 mm)