1 How Dietary Deficiency Studies Have Illuminated the Many Roles …
11
progenitor cells to differentiate into neurons in a dish, it was reasonable to speculate
that vitamin A might be required to sustain neurogenesis in the regenerative olfactory
system throughout adult life. This was speculative because at the time, it was not
known whether the postnatal olfactory system actively metabolized or responded to
vitamin A or its derivatives. Together, my lab group of undergraduate and graduate
students, research assistants, and a postdoctoral fellow showed that cellular retinoic
acid binding proteins (Asson-Batres et al. 2003a, b), retinaldehyde dehydrogenases
(Asson-Batres and Smith 2006), and vitamin A and retinoic acid (Asson-Batres et al.
2009) are present in the postnatal rodent olfactory system. Using well-characterized
antibodies directed against RARα and RARγ (Rochette-Egly et al. 1991; Gaub et al.
1989), we found that RARγ is present in both immature (GAP 43
+ ) and mature
(OMP
+ ) olfactory neurons (Fig. 1.2); we did not detect RARα protein expression
in postnatal mouse sections. We did not evaluate RARβ protein expression because
validated, specific, antibodies were not available.
With knowledge that postnatal rodent olfactory tissue had a potentially active
vitamin A responsive system in place, we set up experiments to test the effects of
dietary vitamin A deficiency on this system. These experiments were based on wellestablished protocols that were developed and improved by many laboratories over
the course of vitamin A research history.
Custom Diets
The vitamin A deficiency research model is reliant upon feeding experimental animals a specially formulated, purified, diet that lacks all sources of vitamin A and
comparing the results with what is observed when animals are fed the same diet
supplemented with a source of vitamin A. Sources of vitamin A include pre-formed
vitamin A (e.g., retinyl acetate, retinyl palmitate) or vitamin A itself (retinol). By
convention, dietary vitamin A content is expressed in international units (IU), with
one IU vitamin A equivalent to 0.3 ug retinol, 0.344 ug retinyl acetate, or 0.55 g
retinyl palmitate (Ross 2010).
The American Institute of Nutrition recommends 4000 IU vitamin A/kg diet
(Reeves et al. 1993), but even so, different amounts of vitamin A equivalents are
found in regular chow maintenance and breeding diets and custom-prepared control
diets (e.g., 4000–22,000 IU vitamin A/kg rodent diet, Table 1.1). Supplementation
with as much as 500,000 IU vitamin A or greater has been used to assess effects of
excess vitamin A intake. There is a positive correlation between the amount of vitamin
A in the diet and the amount present in the liver, whereas circulating levels of vitamin
A are fairly constant (Table 1.2). The observation that plasma/serum vitamin A is
maintained at a steady state suggests circulatory levels are regulated in coordination
with the ever changing systemic demands of extrahepatic tissues. Olson concluded
from his studies that plasma/serum vitamin A levels are “homeostatically controlled
over the physiologic range of liver vitamin A concentrations” (Olson 1984).
11
progenitor cells to differentiate into neurons in a dish, it was reasonable to speculate
that vitamin A might be required to sustain neurogenesis in the regenerative olfactory
system throughout adult life. This was speculative because at the time, it was not
known whether the postnatal olfactory system actively metabolized or responded to
vitamin A or its derivatives. Together, my lab group of undergraduate and graduate
students, research assistants, and a postdoctoral fellow showed that cellular retinoic
acid binding proteins (Asson-Batres et al. 2003a, b), retinaldehyde dehydrogenases
(Asson-Batres and Smith 2006), and vitamin A and retinoic acid (Asson-Batres et al.
2009) are present in the postnatal rodent olfactory system. Using well-characterized
antibodies directed against RARα and RARγ (Rochette-Egly et al. 1991; Gaub et al.
1989), we found that RARγ is present in both immature (GAP 43
+ ) and mature
(OMP
+ ) olfactory neurons (Fig. 1.2); we did not detect RARα protein expression
in postnatal mouse sections. We did not evaluate RARβ protein expression because
validated, specific, antibodies were not available.
With knowledge that postnatal rodent olfactory tissue had a potentially active
vitamin A responsive system in place, we set up experiments to test the effects of
dietary vitamin A deficiency on this system. These experiments were based on wellestablished protocols that were developed and improved by many laboratories over
the course of vitamin A research history.
Custom Diets
The vitamin A deficiency research model is reliant upon feeding experimental animals a specially formulated, purified, diet that lacks all sources of vitamin A and
comparing the results with what is observed when animals are fed the same diet
supplemented with a source of vitamin A. Sources of vitamin A include pre-formed
vitamin A (e.g., retinyl acetate, retinyl palmitate) or vitamin A itself (retinol). By
convention, dietary vitamin A content is expressed in international units (IU), with
one IU vitamin A equivalent to 0.3 ug retinol, 0.344 ug retinyl acetate, or 0.55 g
retinyl palmitate (Ross 2010).
The American Institute of Nutrition recommends 4000 IU vitamin A/kg diet
(Reeves et al. 1993), but even so, different amounts of vitamin A equivalents are
found in regular chow maintenance and breeding diets and custom-prepared control
diets (e.g., 4000–22,000 IU vitamin A/kg rodent diet, Table 1.1). Supplementation
with as much as 500,000 IU vitamin A or greater has been used to assess effects of
excess vitamin A intake. There is a positive correlation between the amount of vitamin
A in the diet and the amount present in the liver, whereas circulating levels of vitamin
A are fairly constant (Table 1.2). The observation that plasma/serum vitamin A is
maintained at a steady state suggests circulatory levels are regulated in coordination
with the ever changing systemic demands of extrahepatic tissues. Olson concluded
from his studies that plasma/serum vitamin A levels are “homeostatically controlled
over the physiologic range of liver vitamin A concentrations” (Olson 1984).