Dietary long-chain polyunsaturated fatty acids upregulate expression of FADS3 transcripts.

The fatty acid desaturase (FADS) gene family at 11q12-13.1 includes FADS1 and FADS2, both known to mediate biosynthesis of omega-3 and omega-6 long-chain polyunsaturated fatty acids (LCPUFA). FADS3 is a putative desaturase due to its sequence similarity with FADS1 and FADS2, but its function is unknown. We have previously described 7 FADS3 alternative transcripts (AT) and 1 FADS2 AT conserved across multiple species. This study examined the effect of dietary LCPUFA levels on liver FADS gene expression in vivo and in vitro, evaluated by qRT-PCR. Fourteen baboon neonates were randomized to three diet groups for their first 12 weeks of life, C: Control, no LCPUFA, L: 0.33% docosahexaenoic acid (DHA)/0.67% arachidonic acid (ARA) (w/w); and L3: 1.00% DHA/0.67% ARA (w/w). Liver FADS1 and both FADS2 transcripts were downregulated by at least 50% in the L3 group compared to controls. In contrast, FADS3 AT were upregulated (L3 > C), with four transcripts significantly upregulated by 40% or more. However, there was no evidence for a shift in liver fatty acids to coincide with increased FADS3 expression. Significant upregulation of FADS3 AT was also observed in human liver-derived HepG2 cells after DHA or ARA treatment. The PPARγ antagonist GW9662 prevented FADS3 upregulation, while downregulation of FADS1 and FADS2 was unaffected. Thus, FADS3 AT were directly upregulated by LCPUFA by a PPARγ-dependent mechanism unrelated to regulation of other desaturases. This opposing pattern and mechanism of regulation suggests a dissimilar function for FADS3 AT compared to other FADS gene products.

Dietary docosahexaenoic acid but not arachidonic acid influences central nervous system fatty acid status in baboon neonates.

The influence of dietary docosahexaenoic acid (DHA, 22:6n-3) and arachidonic acid (AA, 20:4n-6) on infant central nervous system (CNS) composition has implications for neural development, including vision, cognition, and motor function. We consider here combined results of three published studies of DHA/AA-containing formulas and breastfeeding to evaluate the CNS tissue response of baboon neonates with varied concentration and duration of DHA/AA consumption [G.Y. Diau, A.T. Hsieh, E.A. Sarkadi-Nagy, V. Wijendran, P.W. Nathanielsz, J.T. Brenna, The influence of long chain polyunsaturate supplementation on docosahexaenoic acid and arachidonic acid in baboon neonate central nervous system, BMC Med. 3 (2005) 11; A.T. Hsieh, J.C. Anthony, D.A. Diersen-Schade, et al., The influence of moderate and high dietary long chain polyunsaturated fatty acids (LCPUFA) on baboon neonate tissue fatty acids, Pediatr. Res. 61 (2007) 537-45; E. Sarkadi-Nagy, V. Wijendran, G.Y. Diau, et al., The influence of prematurity and long chain polyunsaturate supplementation in 4-week adjusted age baboon neonate brain and related tissues, Pediatr. Res. 54 (2003) 244-252]. A total of 43 neonates born spontaneously at term, or preterm by Cesarean section, consumed diets with DHA-AA (%w/w) at several levels: none (0,0), moderate (0.3, 0.6), or high (>0.6, 0.67 or 1.2). CNS fatty acids were analyzed at 4 and 12 weeks postpartum for term baboons and 7.5 weeks for preterm neonates. CNS DHA was consistently greater by 5-30% in neonates consuming DHA and nearer 30% for cortex. In contrast, CNS AA was unaffected by dietary AA and decreased in all structures with age. Dietary DHA consistently supports greater CNS DHA and maintenance of cortex DHA concentration with feeding duration, while CNS AA is not related to dietary supply. These data on structure-specific LCPUFA accretion may provide insight into neural mechanisms responsible for suboptimal functional outcomes in infants consuming diets that do not support the highest tissue DHA levels.

Differential cerebral cortex transcriptomes of baboon neonates consuming moderate and high docosahexaenoic acid formulas.

BACKGROUND: Docosahexaenoic acid (DHA, 22:6n-3) and arachidonic acid (ARA, 20:4n-6) are the major long chain polyunsaturated fatty acids (LCPUFA) of the central nervous system (CNS). These nutrients are present in most infant formulas at modest levels, intended to support visual and neural development. There are no investigations in primates of the biological consequences of dietary DHA at levels above those present in formulas but within normal breastmilk levels. METHODS AND FINDINGS: Twelve baboons were divided into three formula groups: Control, with no DHA-ARA; "L", LCPUFA, with 0.33%DHA-0.67%ARA; "L3", LCPUFA, with 1.00%DHA-0.67%ARA. All the samples are from the precentral gyrus of cerebral cortex brain regions. At 12 weeks of age, changes in gene expression were detected in 1,108 of 54,000 probe sets (2.05%), with most showing <2-fold change. Gene ontology analysis assigns them to diverse biological functions, notably lipid metabolism and transport, G-protein and signal transduction, development, visual perception, cytoskeleton, peptidases, stress response, transcription regulation, and 400 transcripts having no defined function. PLA2G6, a phospholipase recently associated with infantile neuroaxonal dystrophy, was downregulated in both LCPUFA groups. ELOVL5, a PUFA elongase, was the only LCPUFA biosynthetic enzyme that was differentially expressed. Mitochondrial fatty acid carrier, CPT2, was among several genes associated with mitochondrial fatty acid oxidation to be downregulated by high DHA, while the mitochondrial proton carrier, UCP2, was upregulated. TIMM8A, also known as deafness/dystonia peptide 1, was among several differentially expressed neural development genes. LUM and TIMP3, associated with corneal structure and age-related macular degeneration, respectively, were among visual perception genes influenced by LCPUFA. TIA1, a silencer of COX2 gene translation, is upregulated by high DHA. Ingenuity pathway analysis identified a highly significant nervous system network, with epidermal growth factor receptor (EGFR) as the outstanding interaction partner. CONCLUSIONS: These data indicate that LCPUFA concentrations within the normal range of human breastmilk induce global changes in gene expression across a wide array of processes, in addition to changes in visual and neural function normally associated with formula LCPUFA.

The influence of moderate and high dietary long chain polyunsaturated fatty acids (LCPUFA) on baboon neonate tissue fatty acids.

Docosahexaenoic acid (DHA) and arachidonic acid (ARA) are now common ingredients in commercial infant formulas, however, the optimal levels have not been established. Our previous data showed that the current amount of DHA in U.S. term formulas, 0.3%w/w, is insufficient to normalize cerebral cortex DHA to levels in breastfed baboon neonate controls (Diau et al.: BMC Medicine 3: 11, 2005). Here, we report on the influence of higher formula DHA levels on 12-wk-old full-term baboon CNS and visceral organs. Fourteen nursery-reared baboons were randomized to one of three diets: control (C, no DHA-ARA); moderate LCPUFA (L, 0.33%DHA-0.67%ARA); high LCPUFA (L3, 1.00%DHA-0.67%ARA). DHA increased significantly in liver, heart, and plasma (all C < L < L3), RBC (C < L, L3), and CNS regions: precentral gyrus (C < L < L3), frontal cortex, inferior and superior colliculi, globus pallidus, and caudate (all C < L, L3). These data extend previous observations indicating that 1) tissue DHA is more sensitive to diet than ARA; 2) cerebral cortex DHA increases with higher levels of DHA than in present commercial formulas; and 3) basal ganglia and limbic system DHA saturate with levels of DHA currently available in formulas. These results imply that higher levels of DHA are necessary to normalize cortex DHA to those found in breastfed animals.

The influence of long chain polyunsaturate supplementation on docosahexaenoic acid and arachidonic acid in baboon neonate central nervous system.

BACKGROUND: Docosahexaenoic acid (DHA) and arachidonic acid (ARA) are major components of the cerebral cortex and visual system, where they play a critical role in neural development. We quantitatively mapped fatty acids in 26 regions of the four-week-old breastfed baboon CNS, and studied the influence of dietary DHA and ARA supplementation and prematurity on CNS DHA and ARA concentrations. METHODS: Baboons were randomized into a breastfed (B) and four formula-fed groups: term, no DHA/ARA (T-); term, DHA/ARA supplemented (T+); preterm, no DHA/ARA (P-); preterm and DHA/ARA supplemented (P+). At four weeks adjusted age, brains were dissected and total fatty acids analyzed by gas chromatography and mass spectrometry. RESULTS: DHA and ARA are rich in many more structures than previously reported. They are most concentrated in structures local to the brain stem and diencephalon, particularly the basal ganglia, limbic regions, thalamus and midbrain, and comparatively lower in white matter. Dietary supplementation increased DHA in all structures but had little influence on ARA concentrations. Supplementation restored DHA concentrations to levels of breastfed neonates in all regions except the cerebral cortex and cerebellum. Prematurity per se did not exert a strong influence on DHA or ARA concentrations. CONCLUSION: 1) DHA and ARA are found in high concentration throughout the primate CNS, particularly in gray matter such as basal ganglia; 2) DHA concentrations drop across most CNS structures in neonates consuming formulas with no DHA, but ARA levels are relatively immune to ARA in the diet; 3) supplementation of infant formula is effective at restoring DHA concentration in structures other than the cerebral cortex. These results will be useful as a guide to future investigations of CNS function in the absence of dietary DHA and ARA.

Formula feeding potentiates docosahexaenoic and arachidonic acid biosynthesis in term and preterm baboon neonates.

Infant formulas supplemented with docosahexaenoic acid (DHA) and arachidonic acid (ARA) are now available in the United States; however, little is known about the factors that affect biosynthesis. Baboon neonates were assigned to one of four treatments: term, breast-fed; term, formula-fed; preterm (155 of 182 days gestation), formula-fed; and preterm, formula+DHA/ARA-fed. Standard formula had no DHA/ARA; supplemented formula had 0.61%wt DHA (0.3% of calories) and 1.21%wt ARA (0.6% of calories), and baboon breast milk contained 0.68 +/- 0.22%wt DHA and 0.62 +/- 0.12%wt ARA. At 14 days adjusted age, neonates received a combined oral dose of [U-13C]alpha-linolenic acid (LNA*) and [U-13C]linoleic acid (LA*), and tissues were analyzed 14 days after dose. Brain accretion of linolenic acid-derived DHA was approximately 3-fold greater for the formula groups than for the breast-fed group, and dietary DHA partially attenuated excess DHA synthesis among preterms. A similar, significant pattern was found in other organs. Brain linoleic acid-derived ARA accretion was significantly greater in the unsupplemented term group but not in the preterm groups compared with the breast-fed group. These data show that formula potentiates the biosynthesis/accretion of DHA/ARA in term and preterm neonates compared with breast-fed neonates and that the inclusion of DHA/ARA in preterm formula partially restores DHA/ARA biosynthesis to lower, breast-fed levels. Current formula DHA concentrations are inadequate to normalize long-chain polyunsaturated fatty acids synthesis to that of breast-fed levels.

Influence of dietary long-chain PUFA on premature baboon lung FA and dipalmitoyl PC composition.

One of the major survival challenges of premature birth is production of lung surfactant. The lipid component of surfactant, dipalmitoyl PC (DPPC), increases in concentration in the period before normal term birth via a net shift in FA composition away from unsaturates. We investigated the influence of dietary DHA and arachidonic acid (AA) on lung FA composition and DPPC concentration in term and preterm baboons. Pregnant animals/neonates were randomized to one of four groups: breast-fed (B), term formula-fed (T-, preterm formula-fed (P-, and preterm fed formula supplemented with DHA-AA (P+). Breast milk contained 0.68%wt DHA and the P+ group formula contained 0.61%wt DHA. In the preterm groups (P- and P+), pregnant females received a course of antenatal corticosteroids. At the adjusted age of 4 wk, neonate lung tissue was harvested, and FA composition and DPPC were analyzed. Palmitate was approximately 28%wt of lung total FA and no significant differences were found among the four treatment groups. In contrast, DPPC in the B group lung tissue was significantly greater than DPPC in the unsupplemented groups, but not compared with the P+ group. The B and P+ groups were not significantly different in DHA and AA, but were different compared with the unsupplemented (T, P-) groups. These results indicate that LCP supplementation increases lung DHA and AA, without compromising overall lung 16:0 or DPPC. The shift in FA composition toward greater unsaturation in the groups consuming LCP supported improved surfactant lipid concentration in preterm neonate lungs.

The influence of prematurity and long chain polyunsaturate supplementation in 4-week adjusted age baboon neonate brain and related tissues.

Clinical studies show that docosahexaenoic acid (DHA) and arachidonic acid (ARA) supplemented formula improve visual function in preterm infants, however improved fatty acid status is known only for plasma and red blood cells (RBC) since target organs cannot be sampled from humans. Baboons were randomized to one of four groups: Term breast-fed (B); Term formula-fed (T-); Preterm formula-fed (P-); and Preterm DHA/ARA-supplemented formula-fed (P+). The P+ contained 0.61 +/- 0.03% DHA and 1.21 +/- 0.09% ARA, and breast milk had 0.68 +/- 0.22% and 0.62 +/- 0.12% as DHA and ARA, respectively. The B and P+ groups had significantly higher DHA concentration in all tissues than T- and P-. The P- group showed dramatically lower DHA content of 35%, 27%, 66%, and 75% in the brain, retina, liver, and plasma, respectively, compared with B. Supplementation prevented declines in DHA levels in the retina, and liver, and attenuated the decline in brain, plasma and RBC of preterm animals. In contrast, ARA was not significantly lower compared with B in any group in any tissue but was significantly elevated in liver and brain. RBC and plasma DHA were correlated with DHA in tissues; RBC/plasma ARA were uncorrelated with tissue ARA. We conclude that 1) DHA drops precipitously in term and preterm primates consuming formula without long chain polyunsaturates, while 22:5n-6 concentration rises; 2) tissue ARA levels are insensitive to dietary LCP supplementation or prematurity, 3) plasma and RBC levels of ARA are uncorrelated with total ARA levels; 4) DHA levels are correlated with group effects and are uncorrelated within groups.