Fetal soft tissue examination

Fetal Soft Tissue Examination by Serial Sectioning... 

Historical control data for embryo-fetal studies at Ricerca Biosciences with CrI OFA SD Rat fetal fixed soft tissue examination... 

Afrer the caesarean examination, all live fetuses are given a soft tissue examination and sexed. Following this, the heads of approximately half of the fetuses in each litter are removed and frxed for subsequent examination by serial sectioning (see Chapter 19). The eviscerated fetal carcasses are individually identified by the dam number and fetus number written on a card tag tied around the waist. 

This chapter descrihes the method used for serial sectioning and soft tissue examination of the Bouin s fixed fetus (mainly whole rat and or mouse fetuses or rabbit fetal heads) for the assessment of developmental and structural abnormalities. Fetuses are examined externally, together with the internal structures of the head as well as the thoracic and abdominal organs. 

Key words Serial sectioning, Fetal soft tissue, Bouin s, Wilson technique. Examination, Abnormalities... 

The fetuses are then examined. Full details of fetal examination are provided in other chapters of this book some techniques are performed on fresh fetuses, while other techniques require prior fixation of the fetuses. For rodents, one half of the fetuses in each litter should be examined for soft tissue (visceral) changes the remainder should be examined for skeletal changes. Rabbit fetuses should be examined for both soft tissue and skeletal abnormalities the heads from half of the fetuses should be examined by serial sectioning, with skeletal examination conducted on the remainder of the fetus. 

This study is performed in two species, a rodent (usually the rat) and a non-rodent (nearly always the rabbit). Pregnant females are fed the treated diet from the day of implantation of the embryo through to the end of gestation (i.e., through the embryonic and fetal periods). The females are euthanized shortly before they would have given birth and the femses are examined to detect any strucmral abnormalities. The dams unfortunately cannot be left to give birth namrally because they tend to eat any malformed pups before they can be examined. The fetuses are sampled and processed for detailed examinations of the soft tissues and the skeleton (see Chapters 16-21). 

Approximately one half of each litter is eviscerated and the eviscerated fetal carcasses are fixed and processed for skeletal examination. The skeletal examinations are performed following maceration of the soft tissues with aqueous potassium hydroxide, staining of the skeleton with Alizarin red, and then passage into glycerol. 

This technique is required for any embryo-fetal developmental toxicity in the rat, mouse, rabbit, or minipig. Caesarean sections are performed on dams before birth. Fetuses are removed and examined externally. Further examinations are then performed to examine internal soft tissues (see Chapters 19 and 20) and skeleton. Fetuses intended for skeletal examinations are eviscerated and the carcasses are fixed. The Alizarin Red S staining process requires several days. The meticulous examination involves the examination of each fetal in a specific order starting from the head to the tail, with the fetus in ventral and dorsal positions. 

This chapter describes methods for the examination of fetal abdominal and thoracic soft tissues by microdissection on either fresh (non-rodent) or fixed (rodent) specimens in order to detect structural abnormalities. With hundreds of fetuses examined for each species (rodent and non-rodent) in regulatory reproductive toxicity assessments (ICH, http //www.ich.org/fileadmin/Public Web Site/ICH ... 

Key words Fetal examination, Visceral, Fresh soft tissue. Fixed tissue. Microdissection... 

The test is terminated by performing a C-section on the day before normal delivery is expected. The uterus is examined for implantation and resorption sites and for live and dead fetuses, and the ovaries are examined for corpora lutea. In rodent studies, half of the fetuses are examined for soft tissue malformations, and the remaining are examined for skeletal malformations. In nonrodents, all fetuses are examined for both soft tissue and skeletal malformations. The various end points that may be examined include maternal toxicity, embryo-fetal toxicity, external malformations, and soft tissue and skeletal malformations. 

In contrast to primates, which abort dead embryos, dead rodent embryos are resorbed and the implantation site is recorded as a resorption site. The numbers of living and dead fetuses, and the number of resorption sites, are counted, and fetal weight, sex, and external malformations are recorded. Fetuses can either be inspected fresh in evaluation of internal soft tissues (known as the Staples technique) or can be placed in a fixative (usually Bouin s solution) and sectioned at a later time (Wilson technique). Other fetuses are fixed in ethanol, cleared in potassium hydroxide, and the cartilage and bone are stained with Alcian Blue and Alizarin Red, respectively. The US FDA recommends that one-third of the rodent fetuses be subjected to visceral examination and that two-thirds be studied for abnormalities of cartilage and bone. The US Environmental Protection Agency (EPA) recommends that one-third to one-half of each litter be examined for skeletal anomalies. For rabbits, all fetuses are to be examined for both visceral and skeletal malformations. 

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