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Plasma and dietary cholesterol in infancy: Effects of early low or moderate dietary cholesterol intake on subsequent response to increased dietary cholesterol
Plasma and Dietary Cholesterol in Infancy : Effects of Early Low or Moderate Dietary Cholesterol Intake on Subsequent Response to Increased Dietary Cholesterol By C. J. Glueck,
R. Tsang,
Variations in dietary cholesterol and polyunsaturated or saturated fatty acids were studied in the first yr of life in 21 normal and 18 hypercholesterolemic neonates to determine if the amount of dietary cholesterol in early infancy affects subsequent adaptation to increased dietary cholesterol in later
W. Balistreri.
and R. Fallat
infancy and childhood. At age 12 mo, plasma cholesterol levels in normal (129 mg/lOO ml) and hypercholesterolemit infants (200 mg/lOO ml) did not appear to be influenced by antecedent low or moderate cholesterol intake in early infancy.
T
0 AIM AT LIFELONG NORMALIZATION of plasma cholesterol levels and potential primary prevention of atherosclerosis in normal and hypercholesterolemic infants, intake of cholesterol and polyunsaturated/saturated fatty acids might be modified in infancy. In infants with neonatal familial Type II hyperlipoproteinemia, plasma cholesterol levels can be normalized in the first yr of life by cholesterol-poor polyunsaturate-rich diets.’ Plasma cholesterol levels are substantially higher in normal or hypercholesterolemic infants receiving breast or cow‘s milk’-’ than in infants taking formula milks with skim milk (polyunsaturate-rich) vegetable oil contents. Approximately 80% of normal infants are being fed relatively low cholesterol, polyunsaturaterich commercial formulae.* Low cholesterol intakes have been evaluated in many infants with familial Type II (l/z %-I% of unselected live births).’ Within this frame of reference, concern has been expressed as to the desirability of cholesterol-poor polyunsaturate-rich diets in early infancy, especially in normal children. It has been postulated that adequate dietary “cholesterol challenge” during early infancy may be required to stimulate development of From
the Departments of Medicine, Fels Division of Pediatric Research, Children’s Research Foundation, and the General Clinical Research Center, University of Cincinnati, College of Medicine, Cincinnati, Ohio. Received for publication May 24, 7972. Supporfed in purf by NIH Grunt 1 ROI HE 14597-01 (Dr. Glueck and Dr. Tsung); by fhe Fels Institute for Developmental Research, Yellow Springs, Ohio (Dr. Tsang); and by General Clinical Research Center Grunt RR-00068-10. A portion of this work was done during Dr. Glue&s tenure us an Established Invesfigufor of the American Heart Association, 1971-1976. C. J. Check, M.D.: ASsociufe Professor of Medicine, Director, Lipoprotein Research Laboratory, and Project Director, General Clinical Research Center, University of Cincinnati, College of Medicine, Cincinnati, Ohio. R. Tsang, M.D.: Assistant Professor of Pediatrics and Obstetrics and Gynecology, Fels Division of Pediatric Research, University of Cincinnati, College of Medicine, Cincinnati, Ohio. W. Balistreri, M.D.: Resident in Pediatrics, University of Cincinnati, College of Medicine, Cincinnati, Ohio. R. Fallat, M.D. : Fellow in Endocrinology and Lipoprotein Research, University of Cincinnati, College of Medicine, Cincinnati, Ohio. Hospital
Metabolism,
Vol. 21, No. 12 (December),
1972
1181
1182
GLUECK
ET AL.
enzyme systems responsible for cholesterol catabolism.*-lo Data of Reiser and Kubat in weanling pigs and rats suggested that animals with low cholesterol intake in early infancy, and high cholesterol intake in later infancy eventually develop higher plasma cholesterol levels than comparable animals, which had continuously received a high cholesterol intake beginning in early infancy.g*10 In this study, variations in dietary cholesterol and polyunsaturates and saturates were studied in early infancy in attempts to determine if early moderate dietary cholesterol intake is necessary for adaptation to increased dietary cholesterol in later infancy. Studies in 21 normal and 18 hypercholesterolemic neonates suggested that at least in the first yr of life, the level of dietary cholesterol intake in early infancy does not appear to affect the subsequent response to increased dietary cholesterol in later infancy. MATERIALS
AND
METHODS
Cord blood screening of unselected consecutive live births at the Cincinnati General Hospital provided a large pool of neonates with “normal” cord blood cholesterol, <100 mg/lOO ml, and “hypercholesterolemic” neonates (cord blood cholesterol >lOO mg/ 100 rnl.1.7 Sixty-five of 1600 unselected neonates screened (3.6%) had cord blood cholesterol >100 mg/lOO ml. In a &mo sampling period, 21 infants who had previously been shown Normal Infunfs: to have “normal” cord blood cholesterol levels were examined. At 11-1.4 mo of age, these nonfasting infants were seen for well baby followup at the Pediatric Clinic of the Cincinnati General Hospital and in the Lipoprotein Research Clinic of the General Clinical Research Center, and a complete dietary history was recorded. Plasma was drawn at that time for cholesterol determination. Followup measurement of plasma cholesterol was made Hypercholesferolemic Infants: at ages 6 and 12 mo in 18 nonfasting infants selected from a group of infants with cord blood cholesterol >lOOm g/100 ml. Eleven of these hypercholesterolemic infants also could be documented to have neonatal and familial Type II hyperlipoproteinemia by virtue of primary hypercholesterolemia and hyperbeta-lipoproteinemia in one parent. The diagnosis of Type II in the parent was made by paper electrophoresis,ll and measurement of total, alpha- and beta- lipoprotein cholesterol concentrations.la-14 Age-adjusted “suggested” upper normal limits for beta-liproprotein cholesterol of Fredrickson, Levy, and Lees12 were used. The other seven hypercholesterolemic infants were illegitimate with normocholesterolemic mothers, or came from kindred where the mother and professed father had normal plasma cholesterol and beta-lipoprotein cholesterol.
Did Normal Infants: No specific dietary instructions were given to mothers of the normal infants. At the out-patient clinic, a detailed dietary history was recorded including the following: (1) Type of milk initially taken after birth and duration this milk was taken; (2) Type of milk taken later in infancy and age at which change was made to this preparation; (3) Number of eggs eaten per wk, and age when eggs were eaten consistently; (4) Types of milk products taken (cheese, butter, ice cream), and age when substantial amounts of these products were eaten; (5) Types of strained or junior or prepared table meats eaten, and age when a meat or meat product was given at least l/day; (6) Use of lard or animal fats vs. vegetable oils in cooking. Hypercholesferolemic Infants: No specific dietary instructions were given to the mothers of hypercholesterolemic infants during the first 6 mo. At the 6 mo well baby visit in the Lipoprotein Research Clinic of the General Clinical Research Center, complete dietary histories were recorded as above. At repeat evaluation at 12 mo of age, dietary history was again recorded.
DIETARY
CHOLESTEROL
1183
IN INFANCY
Diet Analysis The amount of cholesterol and estimated ratios of polyunsaturated/saturated fatty acids in the infant’s diets were estimated from analysis of known food intake using tables from the Department of Agriculture, handbook No. 817 and data from studies by Woodruff2 Hughes,18 and Hansen.lo.20 Direct measurement of the amount of cholesterol in certain formulae, and a series of strained infant foods was made using a standard Folch extraction.21 By use of dietary history and analysis Selection of Normal Infants by Diet Patterns: of baby foods as above, infants were divided into groups receiving “low” cholesterol or cholesterol intake in the first 6-8 mo of life, with all infants receiving “moderate” cholesterol intake (500 mg/day or >) in the second 6 mo of life. “Low” “moderate” dietary cholesterol in the first 6 mo of life was estimated at 30-50 mg/day, P/S 2.5-3.5/l dietary cholesterol in the first 6 mo of life was estimated at (Tables 1 and 3). “Moderate” 120-200 mg/day, P/S 0.4-0.8/l (Tables 2-4). Thirteen normal infants were selected by intake in the first virtue of “normal” cord blood cholesterol, relatively "low" cholesterol 6-8 mo of life, and availability for resampling at 11-14 mo (Table 1). Eight normal infants intake in the first 6-8 mo of life, and were selected by virtue of “moderate” cholesterol availability for resampling at 11-14 mo (Table 2). An additional criterion for selection involved estimated dietary cholesterol intake of 500 mg/day or more in both groups of infants in the second 6 ma of life. Selection of Hypercholesterolemic Infants by Diet Patterns: Seven hypercholesterolemic infants were selected by virtue of relatively “low” dietary cholesterol intake in the first 6 mo of life, followed by increased dietary cholesterol in the second 6 mo of life (Table 3). Eleven hypercholesterolemic infants were selected by virtue of “moderate” dietary cholesterol intake in the first 6 mo (Table 4). In the second 6 mo, both groups of infants received estimated dietary cholesterol intakes of 500 mg/day or more.
Analytical Methods All plasma samples were extracted with Zeolite and Isopropanol.15 Cholesterol levels in these extracts were then measured with the Technican AutoAnalyzer. Plasma triglycerides were determined by the method of Kessler and Lederer.15 Cholesterol content of strained baby foods (Table 5) was measured in quadruplicate samples from two different lots of standard commercial baby foods, by the Folch extraction.21 Cholesterol content of homogenized milks, evaporated milks, and commercially prepared formulae (similac, modilac) was also measured in quadruplicate from several lots, using the Falch method.21
Statistical Analysis For descriptive purposes, the group of infants that had a relatively “low” cholesterol intake initially and was changed to a “moderate” cholesterol intake subsequently will be termed “law/moderate”, and the group that had a “moderate” cholesterol intake throughout, will be termed “moderate/moderate”. Differences between various groups of infants on the varied diets were analyzed by Student’s t test,22 and by the Wilcoxenden-White nonparametric test for differences.22
RESULTS
Estimation of Dietary Cholesterol In both normal
Content and P/S Ratio
and hypercholesterolemic
groups
of infants
in the first
6 mo of life, the main caloric intake was derived from the various milks and
formula preparations. By use of published tables on cholesterol and polyunsaturated/saturated fatty acid content,2*17-20 and by analysis of cholesterol content of various formulae,2l an estimate of the amount of dietary cholesterol and P/S ratio could be made for the first 6 mo of life. Infants on approximately 24 oz of milk or formula received about 100 mg cholesterol (P/S 0.6/l)
M F M F F F F M M F M F F
JF PH KN TR cc TJ SH TT WR KH DP JA LG (Mean ? 1 SDM)
6 6 8 6 7 7 6 7 8 8 6 6 6 6.7 2 .8
13 14 12 11 12 11 11 12 14 12 14 12.5 11 12.3 2 1.2
Hmg Hmg Hw
Hmg
Hw Hmg
8z!z5
4
Hmg
-
7 7 10 4 5 7 -
Hmg Hmg Hmg Hmg
Smlac Mdlac Mdlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac
Hmg Hmg
7 21 7 -
Milkg
Milk* Eggs11
Dietary
*Approximate dietary cholesterol (birth-6 mo) = 30-50 mg/day; approximate P/S (birth-6 mo) = 2.5-3.5/l. -tApproximate dietary cholesterol (6-12 mo) = 500-1000 mg/day. SMilk preparation in first 6 mo. §Milk preparation in second 6 mo. ‘IAverage egg intake/wk in second 6 mo. Abbreviations: Smlac, similac formula milk: Hmg, homogenized cow’s milk; and Mdlac, modilac formula milk.
Sex
Infant
Age When Diet Cholesterol Increased (mo)
Dietary Cholesterol in the First Six Months of Life and “Moderate’? Second Six Months of Life in Normo-cholesterolemic Neonates
Age of Repeat Sampling (mo)
Table 1. Effects of “Low”’
44 55 52 75 88 78 54 40 64 73 79 64 66 64?14
Cord Blood Cholesterol (mg/100 ml)
Cholesterol
in the
99 102 140 115 158 154 138 168 140 82 153 117 112 129-c26
Plasma Cholesterol at Followup (mg/100 ml)
F
M M
F F M M F
AN
RK CM
TW PC MP GL CD (Mean & 1 SDM)
11 11 13 13 11 12.4 -c 1.3
14 14
12
6 6 6 6 8 6.2 Z!I .7
6 6
6
(mo)
Evap Evap Evap Evap Evap
Evap Evap
Evap
Milk*
Hmg Hmg Hmg Hmg Evap
Hmg Hmg
Hmg
Milkg
Dietary
mo) =
8 3 5 6 6 6&3 0.4-0.8/l.
10-14 7
0
EggsI/
in the First Six Months of Life and Moderatet of Life in Normo-Cholesterolemic Neonates
Increased
Diet Cholesterol
Age When
Dietary Cholesterol Second Six Months
Age of Repeat Sampling (mo)
of Moderate*
*Approximate dietary cholesterol (birth-6 mo) = 120-200 mg/day: approximate P/S (birth-6 tApproximate dietary cholesterol (6-12 mo) = 500-800 mg/day. *Milk preparation in the first six mo. §Milk preparation in the second six mo. /IAverage egg intake/wk in second six mo. Abbreviations: Evap. canned condensed milk formula: and Hmg, homogenized cow’s milk.
Sex
Infant
Table 2. Effects
54 74 60 54 69 64 2 8
75 63
63
Cholesterol
Cord Blood
Cholesterol
in the
150 116 68 135 123 128227
144 141
144
Followup
Plasma Cholesterol at
4
B
f
Z
?
b ;;I
6
F M F F F F F
T’ODT G’W Poll BJ MF HWT SLIT (Mean + 1 SDM)
6 6 6 8 8 6 6 6.6 f 1.0
Smlac Smlac Smlac Smlac Mdlac Smlac Mdlac
Milk*
Smlac Hmg Hmg Evap Hmg Hmg Hmg
Milkg
11
21 3 14 14 7 3 6 9.7 ‘- 6.6
Eggs
101 267 103 114 190 132 177 154.8 + 61 156 -c 697
(mg/lWml)
Cord Cholesterol
Dietary Cholesterol in the First Six Months of Life and “Moderate”t Second Six Months of Life, in Hypercholesterolemic Neonates
Age When Diet Cholesterol Increased (mo)
of “Low””
242 200 222 170 156 115 200 ?I 60 202 r 721
milk.
298
Plasma Cholesterol 12 mo (mg/lOO ml)
-
in the
154 94 122 136 138 129 2 23 131 i- 268
Plasma Cholesterol 6 mo (mg/lW ml)
Cholesterol
cow’s
Dietary
‘Approximate dietary cholesterol (birth-6 mo) = 30-50 mg/day; approximate P/S (birth-6 mo) = 2.5-3.5/l. tApproximate dietary cho!esterol (6-12 mo) = 500-1000 mg/day. *Milk preparation in first 6 mo. @Iilk preparation in second 6 mo. JIAverage egg intake/wk in second 6 mo. rlnfants with documented neonatal and familial Type II. Abbreviations: Smlac, similac; Mdlac, modilac; Evap, canned condensed milk formula; and Hmg, homogenized
Sex
Infant
Table 3. Effects
6 6
6 6 6 6 6.4 St 6
F F
M
M
F M
M F F F
WI T’ODI[
JHB
EB
JE SE
IJ
Evap Evap Evap Evap
Evap Evap
Breast Evap
Evap Evap
Evap
3
4
Ii
2 14 7 5 5.2 -e 3.6
-
Hmg Hmg Hmg Evap Hmg Hmg
-
-
4 3
Eggs
102 101 146 120 116 145 115 115 119 & 14.5 118” 61
111
117 119
Cord Cholesterol (mg/iOO ml)
Dietary
0.4-0.6/l.
144 200 163 167 136 221 193 263 169 2 37 210 ‘- 3or
205
176
Plasma Cholesterol 6 mo (mg/lOO ml)
mo) =
Neonates
of Life and “Moderate’?
of Life in Hypercholesterolemic
in the First Six Months
Evap Hmg
Hmg Hmg Hmg
Milkp
Six Months
Second
Milk*
Choles:erol
Dietary
*Approximate die!ary cholesterol (birth-6 mo) = 120-200 ma/dav; approximate P/S (birth4 tApproximate dietary cholesterol (612 mo) = 500-600 mg/dayy . *Milk preparation in first 6 mo. §Milk preparation in second 6 mo. l/Average egg intake/wk in second 6 mo. IInfants with documented neonatal and familial Type II. Abbreviations: Evap, canned condensed milk formula; and Hmg, homogenized cow’s milk.
EDB D”ll (Mean & 1 SDM)
Jsll
6 6
M
ww
6 6
8
Sex
Infant
of “Moderate”’
Age When Diet Cholesterol Increased (mo)
Table 4. Effects
136 191 162 133 159 230 222 260 197 k 56 225 -c 567
292
204 132
Plasma Cholesterol 12 mo (mg/lOO ml)
Cholesterol
in the
136
274
-
Plasma Cholesterol 18 mo
4
2
z
z
g i-
F
h
GLUECK
1188
ET AL.
from cow’s milk, 150 mg from breast milk (P/S O.&J/l), and 25 mg from skim milk-vegetable oiI formula (P/S 4/1).l The groups of infants were divided (Tables 1 and 3) into those with 30-50 mg cholesterol intake per day during the first 6 mo (low cholesterol formulae) and those taking approximately 120-200 mg cholesterol per day (cow’s or breast milk) (Tables 2 and 4). The amount of dietary cholesterol gradually increased (most noticeably at 6-8 mo as all infants grew older, discontinued skim milk-corn oil formula feeding, took increasing amounts of homogenized milks, “baby foods”, prepared “table foods”, and particularly eggs. The measured cholesterol content of selected baby foods is summarized in Table 5. Chicken had a moderately higher cholesterol content than expected, but this was reproducible throughout all assays. The amount of cholesterol in the diet was reestimated in all infants at age of resampling (11-14 mo), and was judged to be moderate if 500 mg cholesterol per day or more was ingested (Tables 1-4). All normal and hypercholesterolemic children were receiving at least 500 mg cholesterol/ day at age 12 mo (since this was a criterion for entrance into the study). Some children were receiving 900 mg cholesterol/day from egg intake alone. By these dietary estimations 8 normal and 11 hypercholesterolemic infants received a moderate amount of dietary cholesterol (120-200 mg/day) during the first 6 mo of life (Tables 2 and 4), while 13 normal and 7 hypercholesterolemit infants received much less cholesterol (30-50 mglday) (Tables 1 and 3). The estimated polyunsaturate/saturate ratio of the diet in the moderate cholesterol groups was 0.4-0.8/l, while in the “low” cholesterol group, the estimated P/S ratio was 2.5-3.5/l.’ Beginning at about 6 mo, all normal and hypercholesterolemic infants shifted to diets higher in cholesterol (LOO1000 mg/day) (Tables l-4). It was then possible to compare groups of infants whose original diets in early infancy had been relatively low in cholesterol, with subsequent shift to diets with moderate cholesterol content and infants whose initial and subsequent diets both had moderate amounts of cholesterol. The cholesterol intake in the first 6 mo of life in both the “low” and “moderate” intake groups was quite homogeneous for each group, since almost all cholesterol came from milk. All infants were switched in the second six months to increased dietary cholesterol (“moderate” cholesterol intake), 500-1000 mg/day (Tables 1-4). EfFects
of
Dietary
Cholesterol
on Plasma Cholesterol
Normal Infants: Mean cord blood cholesterol levels in the two groups of normal infants (low/moderate and moderate/moderate) were comparable (64 and 64 mg/lOO mI; Tables I and 2). The change in diet to one of increased cholesterol and decreased polyunsaturates occurred at about 6 mo. Plasma cholesterol levels at ages 11-14 mo (129, 128 mg/lOO ml) were similar in the two diet groups, without evidence for higher levels in those infants whose diets had shifted from early “low cholesterol” intake to later “moderate” cholesterol intake (Tables 1 and 2). HyperchoZesteroIemic Infants: In the hypercholesterolemic children, early feeding of a diet “low” in cholesterol had no evident effect on the subsequent
DIETARY
CHOLESTEROL
1189
IN INFANCY
response to diet higher in cholesterol (Tables 3 and 4). At age 6 mo, infants on the low cholesterol polyunsaturate-rich diets had much lower plasma cholesterol levels (129*22 mg/lOO ml) (Table 3) as compared to infants maintained from birth on “moderate” cholesterol intake (194%5) (Table 4). The change in diet to one of increased cholesterol and decreased polyunsaturates occurred at approximately 6 mo of age. Reevaluation of plasma cholesterol at 12 mo of age showed comparable levels for both (low/moderate, moderate/moderate) dietary groups, with mean levels in the 197 to 200 mg/ 100 ml range. Irrespective of dietary cholesterol intake in the first 6 mo, comparable and substantial hypercholesterolemia was evident in both dietary groups by 12 mo of age. Hypercholesterolemic Infants with Documented Familial Type 11: Five infants in the low/moderate dietary group had proven familial Type II (Table 3). For these five infants, mean (‘SD) cord blood cholesterol, plasma cholesterol at 6 and 12 mo were respectively: 156*69, 131226, and 202272 mg/lOO ml, while mean values in the two other infants were respectively: 152, 122, and 196 mg/lOO ml. Six infants in the moderate/moderate dietary group had proven familial type II (Table 4). Mean cord blood cholesterol for these six Type II infants (118 mg/100 ml) and for the other five hypercholesterolemic neonates (122 mg/100 ml) were similar. Plasma cholesterol at 6 mo with familial Type II infants was 210*30, and was 225259 at 12 mo. Mean plasma cholesterol at ages 6 and 12 mo in the remaining five infants (158*21; 163*30 mg/lOO ml) was considerably lower than in familial Type II, but showed the same pattern of response to diet. Males vs. Females: In view of the reported sex-related difference in plasma cholesterol in infancy 8 effects of diet on plasma cholesterol were compared in males and females (Table 6). No significant differences in cord blood cholesterol, or plasma cholesterol at ages 6 and 12 mo were perceived. DISCUSSION
Although the exact type of diet was not prescribed for these infants studied, selection of infants taking either cow or breast milk or vegetable oil-skim milk formula allowed a comparison of two dietary groups: moderate/moderate and low/moderate dietary groups for normal and hypercholesterolemic infants. In both normal and hypercholesterolemic infants, the gradual increase in dietary cholesterol and saturated fatty acids in the first yr of life is accompanied by an increase from mean cord blood choIesterol levels of 64 mg/ Table 5. Cholesterol
Content of Strained Baby Food (mg/lOO g)
Food
Cholesterol
Food
Cholesterol
Egg Yolk Liver
352 300
Beef Ham
60 60
Vanilla Custard
38
Pudding’ Chicken
130 102
Veal Lamb
80 60
Cottage Cheese Peast
27
‘With eggs. tCooked with butter.
Food
Cholesterol
16
1190
GLUECK
ET AL.
100 ml (for normals) or 119-129 mg/lOO ml (for hypercholesterolemic infants) to levels of 128-129 mg/lOO ml for normals (at age 12 mo) and 197-200 mg/ 100 ml for hypercholesterolemic infants (at age 12 mo). This gradual increase in plasma cholesterol in normal and hypercholesterolemic neonates did not, under the conditions of this study, appear to be affected by early exposure to low or moderate cholesterol intake followed by ingestion of substantially increased cholesterol in the second 6 mo of life. For infants with proven neonatal and familial Type II, low cholesterol intake in early neonatal periods did not impair subsequent response to increased dietary cholesterol. At 12 mo of age (and after 6 mo of moderate cholesterol intake), familial Type II infants had the highest plasma cholesterol values (mean 225 mg/lOO ml), Comparable results (but on a much shorter study period) were obtained by Lindquist and Malmcrona,5 who gave a corn oil base formula to five infants for 8 days, followed by 2 wk of cream or breast milk. At followup, total lipids and plasma cholesterol in infants progressing from corn oil to cream-breast milk could not be differentiated from those of children on breast milk throughout. These findings in infants differ appreciabIy from studies in rat&O and pigs9 where it was postulated that cholesterol in the diet of the suckling controls development of enzymes concerned with cholesterol homeostasis in the adult. Kubat et al. weaned rats at 18 and 30 days, then fed a “high fat diet” until 10 mo of age, followed by 2% mo of cholesterol-rich diet. Although there were no group differences in serum cholesterol at 10 mo, serum cholesterol was higher at 12% mo in the early weaned group.lO Reiser fed a small number of pigs low cholesterol (1.5 mg/dl) or moderate cholesterol (114 mg/dl) milk, followed by normal pig feed and serial increases in addi-
Table 6
Neonatal Hypercholesterolemia Comparison of cord blood cholesterol, plasma cholesterol at 6 and 12 mo, males vs. females (moderate/moderate diet group) Males (N=5)
Cord blood cholesterol (mg/lOO ml) Plasma cholesterol, age 6 mo Plasma cholesterol, age 12 mo
111.6 2 9.3 166.3 k 31 165 k-l.6
Females (N=6)
Significance
125.2 -c 15.9 206.6 & 31 223 ‘- 60
Normocholesterolemic Neonates Comparison of cord blood cholesterol, plasma cholesterol at 12 mo, males vs. females (moderate/moderate diet group) Females Males (N=4)
Cord blood cholesterol (mg/lOO ml) Plasma cholesterol, age 12 mo
63 122
-t 8.8 2 36.2
Significance
(N-4)
65 133
k 6.6 s 16.5
Normorcholesterolemic Neonates Comparison of cord blood cholesterol, plasma cholesterol 12 mo, males vs. females (low/moderate diet group)
Cord blood cholesterol (mg/lOO ml) Plasma cholesterol, age 12 mo
NS NS NS
NS NS
at
Males (N=5)
Females (N=8)
55.8 -1- 15.9 140 r+ 25.7
69.1 -c 11.6 122.5 & 25.9
Significance
NS NS
DIETARY
CHOLESTEROL
1191
IN INFANCY
tional exogenous cholesterol. At 17 mo, cholesterol levels were higher in female pigs originally maintained on low cholesterol milk.9 Some of the apparent dichotomy between human and animal data may lie in the timing and duration of both the “suckling period” and the life span. The age of resampling is obviously critical; Kubat found no differences between dietary groups at age IO mo, with appreciable differences at 12% mo. Extended followup on ad lib. diets with 600 mg cholesterol/day (or more) will be necessary in our infant group to ascertain if the effects of “early low cholesterol diets” are not expressed.much later in the child’s life, rather than 5-6 mo after the change in diet. Other reservations as to the role of the “low cholesterol, polyunsaturaterich diet” in normal or hypercholesterolemic children have centered about the findings that in several animal speciesz3 increased levels of unsaturated fat lead to increased requirement for Vitamin E. Moreover, Hassan et al. have described a syndrome involving edema, papular erythema, an elevated platelet count, and anemia in a small series of premature infants receiving formula relatively rich in polyunsaturated fatty acids.2” Dayton et al. have also described in an elderly population lowered plasma Vitamin E and increased red cell hemolysis during a low cholesterol diet, rich in polyunsaturates.25 Athough comparable changes have not been reported in term infants on less radical dietary formulae than that used by Hassan et a1.,“4 approximately 60% of infants on low cholesterol, polyunsaturate-rich formulae in our studies have received additional Vitamin E supplementation, and none (with or without Vitamin E supplementation) developed the lesions or syndrome described above. Against the potential and for the most part still theoretical problems encountered with early low cholesterol polyunsaturate-rich feeding must be weighed the potential benefit of amelioration of hypercholesterolemia in neonatal familial Type II,ls7 and the much lower cholesterol levels in the normocholesterolemic child’-6*8 on low cholesterol diets. It has been suggested that dietary intervention in the infant with familial type II be started at 1 yr of age 26,27 although our experience suggests that modest intervention in the for6 of low cholesterol, polyunsaturate-rich milks can profitably be started even earlier.lf7 For the normal child, prudent restriction in dietary cholesterol intake might potentially be useful in a general reduction in plasma cholesterol levels. Primary prevention of atherosclerosis in normal and hypercholesterolemic children may potentially start in childhood. Long-term (generation) studies in normal and hypercholesterolemic children will need to be carried out to determine the efficacy (or lack of efficacy) of long-term cholesterol lowering regimens in atherosclerosis prevention. Under the restricted, short-term conditions of this study, very early low cholesterol intake does not appear to adversely affect subsequent ability to maintain “normal cholesterol homeostasis” when cholesterol intake increases in later infancy. REFERENCES 1. Glueck, C. J., and Tsang, R. C.: Neonatal Familial Type II Hyperlipoproteinemia: Effects of diet on plasma cholesterol
in the first year of life. Amer. 25 :224,
1972.
J. Clin. Nutr.
1192 2. Woodruff, C. W., Bailey, M. C., Davis, J. T., Rogers, N., and Conglio, J. G.: Serum lipids in breast-fed infants and in infants fed evaporated milk. Amer. J. Clin. Nutr.
14:83,1964. 3.Lowe, C. J., Mosovich,
proteins
in human
serum.
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ET AL.
J. Clin.
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34:1345,1955. 15. Kessler, G., and Lederer, metric Skeggs,
measurement L. T. Jr.
H.: Fluoritriglycerides. In Automation in
of (Ed.):
Analytical Chemistry. New York, Mediad, L. L., and Pessin, V.: Effects of protein level and type of heat 1966, p. 341. 16. Total cholesterol procedure, N-24: In treatment of milk formulas on growth and maturation of infants. J. Pediat. 64:666, AutoAnalyzer Manual, Chauncey, N. Y., Technicon Instruments Corp., 1964. 1964. 4. Pomeranze, J.: Serum cholesterol stud17.Watt, 8. K., and Merrill, A. L.: Comies in infants: A comparison of infants fed position of Foods: Raw, Processed, Prebreast milk, evaporated milk and prepared pared. Agriculture Research Service Handmilk formulas. Amer. J. Clin. Nutr. 9:570, book Number 8, Washington, D.C., U. S. Government Printing Office, 1963. 1961. 5. Lindquist, B., and Malmcrona, R.: 18.Hughes, G., Kelly, V. J., and Stewart, Dietary fat in relation to serum lipids in R. A.: Linoleic acid-an essential nutrient. the normal infant. Amer. J. Dis. Child. Clin. Pediat. 2:555,1963. 99:55,1950. 19.Hansen, A. E., et al.: Role of linoleic 6. Ballester, D., et al.: Serum cholesterol acid in infant nutrition. Pediatrics 31(Suppl. in Chilean newborn infants and its evalua1, Part II) :171, 1963. tion during the first months of life. Helv. 20.-, et al.: The relation of linoleic acid Pediat. Acta 2:227,1965. to infant feeding. Acta Paediat. ~l(Suppl. 7.Glueck, C. J., Heckman, F., Schoenfeld, 137):5, 1962. M., Steiner, I’., and Pearce, W.: Neonatal 21.Folch, J., Lees, M., and Sloane-Stanley, Familial Type II Hyperlipoproteinemia: cord G. H.: A simple method for the isolation blood cholesterol in 1800 births. Metabolism and purification of total lipids from animal 20597, 1971. tissues. J. Biol. Chem. 226:497, 1957. 8. Fomon, S. J.: A pediatrician looks at 22.Snedecor, G. W., and Cochran, W. G.: early nutrition. Bull. N.Y. Acad. Med. 47: In Statistical Methods (ed. 6).Ames, Iowa,
569,1971. 9. Reiser,
Iowa R.: Control of adult serum by the nutrition of the suckling: report. Circulation 63(Suppl. II):
State
University
Press,
1967, p. 91,
128. 23. Harris,
cholesterol a progress 11-3, 1971. 10.Kubat, K., Hahn, P., and Koldovsky, 0.: Utilization of Nutrients During Postnatal Development Vol. 33. New York Pergamon Press, 1966,p. 154. 11.Lees, R. S., and Hatch, F. T.: Sharper separation of lipoprotein species by paper electrophoresis in albumin-containing buffer. J. Lab. Clin. Med. 61:518, 1963. 12. Frederickson, D. S., Levy, R. I., and Lees, R. S.: Fat transport in lipoproteins: an integrated approach to mechanisms and disorders. New Eng. J. Med. 276:32, 1967. 13. Fredrickson, D. S., Levy, R. I., and Lindgren, F. T.: A comparison of heritable abnormal lipoprotein patterns as defined by two different techniques. J. Clin. Invest.
P. L., and Embree, N. D.: Quantitative consideration of the effect of polyunsaturated fatty acid content of the diet upon the requirements for Vitamin E. Amer. J. Clin.Nutr. 133385,1963. 24.Hassan,H., Hashim, S. A., VanItallie, T. B., and Segrell, W. H.: Syndrome in premature infants associated with low plasma Vitamin E levels and high polyunsaturated fatty acid diet. Amer. J. Clin. Nutr. 19:147,
47~2446,1968. 14.Havel, R. J., Eder, H. A., and Bragdon,
49:165,1972. 27.Filer, L. J., et al.: Childhood
J. H.: Distribution and chemical composition of ultracentrifugally separated lipo-
coronary
1966. 25.Dayton,S., Hashimoto,
S., Rosenblum, D., and Pearce, M. I..: Vitamin E status of humans during prolonged feeding of unsaturated fats. J. Lab. Clin. Med. 65:739,
1965. 26.Mitchell,
S., Blount, S. G., Blumenthal, S., Jesse, M. J., and Weidman, W. H.: The pediatrician and atherosclerosis. Pediatrics
1972.
heart
disease.
Pediatrics
diet and 49 :308,
R. Tsang,
Variations in dietary cholesterol and polyunsaturated or saturated fatty acids were studied in the first yr of life in 21 normal and 18 hypercholesterolemic neonates to determine if the amount of dietary cholesterol in early infancy affects subsequent adaptation to increased dietary cholesterol in later
W. Balistreri.
and R. Fallat
infancy and childhood. At age 12 mo, plasma cholesterol levels in normal (129 mg/lOO ml) and hypercholesterolemit infants (200 mg/lOO ml) did not appear to be influenced by antecedent low or moderate cholesterol intake in early infancy.
T
0 AIM AT LIFELONG NORMALIZATION of plasma cholesterol levels and potential primary prevention of atherosclerosis in normal and hypercholesterolemic infants, intake of cholesterol and polyunsaturated/saturated fatty acids might be modified in infancy. In infants with neonatal familial Type II hyperlipoproteinemia, plasma cholesterol levels can be normalized in the first yr of life by cholesterol-poor polyunsaturate-rich diets.’ Plasma cholesterol levels are substantially higher in normal or hypercholesterolemic infants receiving breast or cow‘s milk’-’ than in infants taking formula milks with skim milk (polyunsaturate-rich) vegetable oil contents. Approximately 80% of normal infants are being fed relatively low cholesterol, polyunsaturaterich commercial formulae.* Low cholesterol intakes have been evaluated in many infants with familial Type II (l/z %-I% of unselected live births).’ Within this frame of reference, concern has been expressed as to the desirability of cholesterol-poor polyunsaturate-rich diets in early infancy, especially in normal children. It has been postulated that adequate dietary “cholesterol challenge” during early infancy may be required to stimulate development of From
the Departments of Medicine, Fels Division of Pediatric Research, Children’s Research Foundation, and the General Clinical Research Center, University of Cincinnati, College of Medicine, Cincinnati, Ohio. Received for publication May 24, 7972. Supporfed in purf by NIH Grunt 1 ROI HE 14597-01 (Dr. Glueck and Dr. Tsung); by fhe Fels Institute for Developmental Research, Yellow Springs, Ohio (Dr. Tsang); and by General Clinical Research Center Grunt RR-00068-10. A portion of this work was done during Dr. Glue&s tenure us an Established Invesfigufor of the American Heart Association, 1971-1976. C. J. Check, M.D.: ASsociufe Professor of Medicine, Director, Lipoprotein Research Laboratory, and Project Director, General Clinical Research Center, University of Cincinnati, College of Medicine, Cincinnati, Ohio. R. Tsang, M.D.: Assistant Professor of Pediatrics and Obstetrics and Gynecology, Fels Division of Pediatric Research, University of Cincinnati, College of Medicine, Cincinnati, Ohio. W. Balistreri, M.D.: Resident in Pediatrics, University of Cincinnati, College of Medicine, Cincinnati, Ohio. R. Fallat, M.D. : Fellow in Endocrinology and Lipoprotein Research, University of Cincinnati, College of Medicine, Cincinnati, Ohio. Hospital
Metabolism,
Vol. 21, No. 12 (December),
1972
1181
1182
GLUECK
ET AL.
enzyme systems responsible for cholesterol catabolism.*-lo Data of Reiser and Kubat in weanling pigs and rats suggested that animals with low cholesterol intake in early infancy, and high cholesterol intake in later infancy eventually develop higher plasma cholesterol levels than comparable animals, which had continuously received a high cholesterol intake beginning in early infancy.g*10 In this study, variations in dietary cholesterol and polyunsaturates and saturates were studied in early infancy in attempts to determine if early moderate dietary cholesterol intake is necessary for adaptation to increased dietary cholesterol in later infancy. Studies in 21 normal and 18 hypercholesterolemic neonates suggested that at least in the first yr of life, the level of dietary cholesterol intake in early infancy does not appear to affect the subsequent response to increased dietary cholesterol in later infancy. MATERIALS
AND
METHODS
Cord blood screening of unselected consecutive live births at the Cincinnati General Hospital provided a large pool of neonates with “normal” cord blood cholesterol, <100 mg/lOO ml, and “hypercholesterolemic” neonates (cord blood cholesterol >lOO mg/ 100 rnl.1.7 Sixty-five of 1600 unselected neonates screened (3.6%) had cord blood cholesterol >100 mg/lOO ml. In a &mo sampling period, 21 infants who had previously been shown Normal Infunfs: to have “normal” cord blood cholesterol levels were examined. At 11-1.4 mo of age, these nonfasting infants were seen for well baby followup at the Pediatric Clinic of the Cincinnati General Hospital and in the Lipoprotein Research Clinic of the General Clinical Research Center, and a complete dietary history was recorded. Plasma was drawn at that time for cholesterol determination. Followup measurement of plasma cholesterol was made Hypercholesferolemic Infants: at ages 6 and 12 mo in 18 nonfasting infants selected from a group of infants with cord blood cholesterol >lOOm g/100 ml. Eleven of these hypercholesterolemic infants also could be documented to have neonatal and familial Type II hyperlipoproteinemia by virtue of primary hypercholesterolemia and hyperbeta-lipoproteinemia in one parent. The diagnosis of Type II in the parent was made by paper electrophoresis,ll and measurement of total, alpha- and beta- lipoprotein cholesterol concentrations.la-14 Age-adjusted “suggested” upper normal limits for beta-liproprotein cholesterol of Fredrickson, Levy, and Lees12 were used. The other seven hypercholesterolemic infants were illegitimate with normocholesterolemic mothers, or came from kindred where the mother and professed father had normal plasma cholesterol and beta-lipoprotein cholesterol.
Did Normal Infants: No specific dietary instructions were given to mothers of the normal infants. At the out-patient clinic, a detailed dietary history was recorded including the following: (1) Type of milk initially taken after birth and duration this milk was taken; (2) Type of milk taken later in infancy and age at which change was made to this preparation; (3) Number of eggs eaten per wk, and age when eggs were eaten consistently; (4) Types of milk products taken (cheese, butter, ice cream), and age when substantial amounts of these products were eaten; (5) Types of strained or junior or prepared table meats eaten, and age when a meat or meat product was given at least l/day; (6) Use of lard or animal fats vs. vegetable oils in cooking. Hypercholesferolemic Infants: No specific dietary instructions were given to the mothers of hypercholesterolemic infants during the first 6 mo. At the 6 mo well baby visit in the Lipoprotein Research Clinic of the General Clinical Research Center, complete dietary histories were recorded as above. At repeat evaluation at 12 mo of age, dietary history was again recorded.
DIETARY
CHOLESTEROL
1183
IN INFANCY
Diet Analysis The amount of cholesterol and estimated ratios of polyunsaturated/saturated fatty acids in the infant’s diets were estimated from analysis of known food intake using tables from the Department of Agriculture, handbook No. 817 and data from studies by Woodruff2 Hughes,18 and Hansen.lo.20 Direct measurement of the amount of cholesterol in certain formulae, and a series of strained infant foods was made using a standard Folch extraction.21 By use of dietary history and analysis Selection of Normal Infants by Diet Patterns: of baby foods as above, infants were divided into groups receiving “low” cholesterol or cholesterol intake in the first 6-8 mo of life, with all infants receiving “moderate” cholesterol intake (500 mg/day or >) in the second 6 mo of life. “Low” “moderate” dietary cholesterol in the first 6 mo of life was estimated at 30-50 mg/day, P/S 2.5-3.5/l dietary cholesterol in the first 6 mo of life was estimated at (Tables 1 and 3). “Moderate” 120-200 mg/day, P/S 0.4-0.8/l (Tables 2-4). Thirteen normal infants were selected by intake in the first virtue of “normal” cord blood cholesterol, relatively "low" cholesterol 6-8 mo of life, and availability for resampling at 11-14 mo (Table 1). Eight normal infants intake in the first 6-8 mo of life, and were selected by virtue of “moderate” cholesterol availability for resampling at 11-14 mo (Table 2). An additional criterion for selection involved estimated dietary cholesterol intake of 500 mg/day or more in both groups of infants in the second 6 ma of life. Selection of Hypercholesterolemic Infants by Diet Patterns: Seven hypercholesterolemic infants were selected by virtue of relatively “low” dietary cholesterol intake in the first 6 mo of life, followed by increased dietary cholesterol in the second 6 mo of life (Table 3). Eleven hypercholesterolemic infants were selected by virtue of “moderate” dietary cholesterol intake in the first 6 mo (Table 4). In the second 6 mo, both groups of infants received estimated dietary cholesterol intakes of 500 mg/day or more.
Analytical Methods All plasma samples were extracted with Zeolite and Isopropanol.15 Cholesterol levels in these extracts were then measured with the Technican AutoAnalyzer. Plasma triglycerides were determined by the method of Kessler and Lederer.15 Cholesterol content of strained baby foods (Table 5) was measured in quadruplicate samples from two different lots of standard commercial baby foods, by the Folch extraction.21 Cholesterol content of homogenized milks, evaporated milks, and commercially prepared formulae (similac, modilac) was also measured in quadruplicate from several lots, using the Falch method.21
Statistical Analysis For descriptive purposes, the group of infants that had a relatively “low” cholesterol intake initially and was changed to a “moderate” cholesterol intake subsequently will be termed “law/moderate”, and the group that had a “moderate” cholesterol intake throughout, will be termed “moderate/moderate”. Differences between various groups of infants on the varied diets were analyzed by Student’s t test,22 and by the Wilcoxenden-White nonparametric test for differences.22
RESULTS
Estimation of Dietary Cholesterol In both normal
Content and P/S Ratio
and hypercholesterolemic
groups
of infants
in the first
6 mo of life, the main caloric intake was derived from the various milks and
formula preparations. By use of published tables on cholesterol and polyunsaturated/saturated fatty acid content,2*17-20 and by analysis of cholesterol content of various formulae,2l an estimate of the amount of dietary cholesterol and P/S ratio could be made for the first 6 mo of life. Infants on approximately 24 oz of milk or formula received about 100 mg cholesterol (P/S 0.6/l)
M F M F F F F M M F M F F
JF PH KN TR cc TJ SH TT WR KH DP JA LG (Mean ? 1 SDM)
6 6 8 6 7 7 6 7 8 8 6 6 6 6.7 2 .8
13 14 12 11 12 11 11 12 14 12 14 12.5 11 12.3 2 1.2
Hmg Hmg Hw
Hmg
Hw Hmg
8z!z5
4
Hmg
-
7 7 10 4 5 7 -
Hmg Hmg Hmg Hmg
Smlac Mdlac Mdlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac Smlac
Hmg Hmg
7 21 7 -
Milkg
Milk* Eggs11
Dietary
*Approximate dietary cholesterol (birth-6 mo) = 30-50 mg/day; approximate P/S (birth-6 mo) = 2.5-3.5/l. -tApproximate dietary cholesterol (6-12 mo) = 500-1000 mg/day. SMilk preparation in first 6 mo. §Milk preparation in second 6 mo. ‘IAverage egg intake/wk in second 6 mo. Abbreviations: Smlac, similac formula milk: Hmg, homogenized cow’s milk; and Mdlac, modilac formula milk.
Sex
Infant
Age When Diet Cholesterol Increased (mo)
Dietary Cholesterol in the First Six Months of Life and “Moderate’? Second Six Months of Life in Normo-cholesterolemic Neonates
Age of Repeat Sampling (mo)
Table 1. Effects of “Low”’
44 55 52 75 88 78 54 40 64 73 79 64 66 64?14
Cord Blood Cholesterol (mg/100 ml)
Cholesterol
in the
99 102 140 115 158 154 138 168 140 82 153 117 112 129-c26
Plasma Cholesterol at Followup (mg/100 ml)
F
M M
F F M M F
AN
RK CM
TW PC MP GL CD (Mean & 1 SDM)
11 11 13 13 11 12.4 -c 1.3
14 14
12
6 6 6 6 8 6.2 Z!I .7
6 6
6
(mo)
Evap Evap Evap Evap Evap
Evap Evap
Evap
Milk*
Hmg Hmg Hmg Hmg Evap
Hmg Hmg
Hmg
Milkg
Dietary
mo) =
8 3 5 6 6 6&3 0.4-0.8/l.
10-14 7
0
EggsI/
in the First Six Months of Life and Moderatet of Life in Normo-Cholesterolemic Neonates
Increased
Diet Cholesterol
Age When
Dietary Cholesterol Second Six Months
Age of Repeat Sampling (mo)
of Moderate*
*Approximate dietary cholesterol (birth-6 mo) = 120-200 mg/day: approximate P/S (birth-6 tApproximate dietary cholesterol (6-12 mo) = 500-800 mg/day. *Milk preparation in the first six mo. §Milk preparation in the second six mo. /IAverage egg intake/wk in second six mo. Abbreviations: Evap. canned condensed milk formula: and Hmg, homogenized cow’s milk.
Sex
Infant
Table 2. Effects
54 74 60 54 69 64 2 8
75 63
63
Cholesterol
Cord Blood
Cholesterol
in the
150 116 68 135 123 128227
144 141
144
Followup
Plasma Cholesterol at
4
B
f
Z
?
b ;;I
6
F M F F F F F
T’ODT G’W Poll BJ MF HWT SLIT (Mean + 1 SDM)
6 6 6 8 8 6 6 6.6 f 1.0
Smlac Smlac Smlac Smlac Mdlac Smlac Mdlac
Milk*
Smlac Hmg Hmg Evap Hmg Hmg Hmg
Milkg
11
21 3 14 14 7 3 6 9.7 ‘- 6.6
Eggs
101 267 103 114 190 132 177 154.8 + 61 156 -c 697
(mg/lWml)
Cord Cholesterol
Dietary Cholesterol in the First Six Months of Life and “Moderate”t Second Six Months of Life, in Hypercholesterolemic Neonates
Age When Diet Cholesterol Increased (mo)
of “Low””
242 200 222 170 156 115 200 ?I 60 202 r 721
milk.
298
Plasma Cholesterol 12 mo (mg/lOO ml)
-
in the
154 94 122 136 138 129 2 23 131 i- 268
Plasma Cholesterol 6 mo (mg/lW ml)
Cholesterol
cow’s
Dietary
‘Approximate dietary cholesterol (birth-6 mo) = 30-50 mg/day; approximate P/S (birth-6 mo) = 2.5-3.5/l. tApproximate dietary cho!esterol (6-12 mo) = 500-1000 mg/day. *Milk preparation in first 6 mo. @Iilk preparation in second 6 mo. JIAverage egg intake/wk in second 6 mo. rlnfants with documented neonatal and familial Type II. Abbreviations: Smlac, similac; Mdlac, modilac; Evap, canned condensed milk formula; and Hmg, homogenized
Sex
Infant
Table 3. Effects
6 6
6 6 6 6 6.4 St 6
F F
M
M
F M
M F F F
WI T’ODI[
JHB
EB
JE SE
IJ
Evap Evap Evap Evap
Evap Evap
Breast Evap
Evap Evap
Evap
3
4
Ii
2 14 7 5 5.2 -e 3.6
-
Hmg Hmg Hmg Evap Hmg Hmg
-
-
4 3
Eggs
102 101 146 120 116 145 115 115 119 & 14.5 118” 61
111
117 119
Cord Cholesterol (mg/iOO ml)
Dietary
0.4-0.6/l.
144 200 163 167 136 221 193 263 169 2 37 210 ‘- 3or
205
176
Plasma Cholesterol 6 mo (mg/lOO ml)
mo) =
Neonates
of Life and “Moderate’?
of Life in Hypercholesterolemic
in the First Six Months
Evap Hmg
Hmg Hmg Hmg
Milkp
Six Months
Second
Milk*
Choles:erol
Dietary
*Approximate die!ary cholesterol (birth-6 mo) = 120-200 ma/dav; approximate P/S (birth4 tApproximate dietary cholesterol (612 mo) = 500-600 mg/dayy . *Milk preparation in first 6 mo. §Milk preparation in second 6 mo. l/Average egg intake/wk in second 6 mo. IInfants with documented neonatal and familial Type II. Abbreviations: Evap, canned condensed milk formula; and Hmg, homogenized cow’s milk.
EDB D”ll (Mean & 1 SDM)
Jsll
6 6
M
ww
6 6
8
Sex
Infant
of “Moderate”’
Age When Diet Cholesterol Increased (mo)
Table 4. Effects
136 191 162 133 159 230 222 260 197 k 56 225 -c 567
292
204 132
Plasma Cholesterol 12 mo (mg/lOO ml)
Cholesterol
in the
136
274
-
Plasma Cholesterol 18 mo
4
2
z
z
g i-
F
h
GLUECK
1188
ET AL.
from cow’s milk, 150 mg from breast milk (P/S O.&J/l), and 25 mg from skim milk-vegetable oiI formula (P/S 4/1).l The groups of infants were divided (Tables 1 and 3) into those with 30-50 mg cholesterol intake per day during the first 6 mo (low cholesterol formulae) and those taking approximately 120-200 mg cholesterol per day (cow’s or breast milk) (Tables 2 and 4). The amount of dietary cholesterol gradually increased (most noticeably at 6-8 mo as all infants grew older, discontinued skim milk-corn oil formula feeding, took increasing amounts of homogenized milks, “baby foods”, prepared “table foods”, and particularly eggs. The measured cholesterol content of selected baby foods is summarized in Table 5. Chicken had a moderately higher cholesterol content than expected, but this was reproducible throughout all assays. The amount of cholesterol in the diet was reestimated in all infants at age of resampling (11-14 mo), and was judged to be moderate if 500 mg cholesterol per day or more was ingested (Tables 1-4). All normal and hypercholesterolemic children were receiving at least 500 mg cholesterol/ day at age 12 mo (since this was a criterion for entrance into the study). Some children were receiving 900 mg cholesterol/day from egg intake alone. By these dietary estimations 8 normal and 11 hypercholesterolemic infants received a moderate amount of dietary cholesterol (120-200 mg/day) during the first 6 mo of life (Tables 2 and 4), while 13 normal and 7 hypercholesterolemit infants received much less cholesterol (30-50 mglday) (Tables 1 and 3). The estimated polyunsaturate/saturate ratio of the diet in the moderate cholesterol groups was 0.4-0.8/l, while in the “low” cholesterol group, the estimated P/S ratio was 2.5-3.5/l.’ Beginning at about 6 mo, all normal and hypercholesterolemic infants shifted to diets higher in cholesterol (LOO1000 mg/day) (Tables l-4). It was then possible to compare groups of infants whose original diets in early infancy had been relatively low in cholesterol, with subsequent shift to diets with moderate cholesterol content and infants whose initial and subsequent diets both had moderate amounts of cholesterol. The cholesterol intake in the first 6 mo of life in both the “low” and “moderate” intake groups was quite homogeneous for each group, since almost all cholesterol came from milk. All infants were switched in the second six months to increased dietary cholesterol (“moderate” cholesterol intake), 500-1000 mg/day (Tables 1-4). EfFects
of
Dietary
Cholesterol
on Plasma Cholesterol
Normal Infants: Mean cord blood cholesterol levels in the two groups of normal infants (low/moderate and moderate/moderate) were comparable (64 and 64 mg/lOO mI; Tables I and 2). The change in diet to one of increased cholesterol and decreased polyunsaturates occurred at about 6 mo. Plasma cholesterol levels at ages 11-14 mo (129, 128 mg/lOO ml) were similar in the two diet groups, without evidence for higher levels in those infants whose diets had shifted from early “low cholesterol” intake to later “moderate” cholesterol intake (Tables 1 and 2). HyperchoZesteroIemic Infants: In the hypercholesterolemic children, early feeding of a diet “low” in cholesterol had no evident effect on the subsequent
DIETARY
CHOLESTEROL
1189
IN INFANCY
response to diet higher in cholesterol (Tables 3 and 4). At age 6 mo, infants on the low cholesterol polyunsaturate-rich diets had much lower plasma cholesterol levels (129*22 mg/lOO ml) (Table 3) as compared to infants maintained from birth on “moderate” cholesterol intake (194%5) (Table 4). The change in diet to one of increased cholesterol and decreased polyunsaturates occurred at approximately 6 mo of age. Reevaluation of plasma cholesterol at 12 mo of age showed comparable levels for both (low/moderate, moderate/moderate) dietary groups, with mean levels in the 197 to 200 mg/ 100 ml range. Irrespective of dietary cholesterol intake in the first 6 mo, comparable and substantial hypercholesterolemia was evident in both dietary groups by 12 mo of age. Hypercholesterolemic Infants with Documented Familial Type 11: Five infants in the low/moderate dietary group had proven familial Type II (Table 3). For these five infants, mean (‘SD) cord blood cholesterol, plasma cholesterol at 6 and 12 mo were respectively: 156*69, 131226, and 202272 mg/lOO ml, while mean values in the two other infants were respectively: 152, 122, and 196 mg/lOO ml. Six infants in the moderate/moderate dietary group had proven familial type II (Table 4). Mean cord blood cholesterol for these six Type II infants (118 mg/100 ml) and for the other five hypercholesterolemic neonates (122 mg/100 ml) were similar. Plasma cholesterol at 6 mo with familial Type II infants was 210*30, and was 225259 at 12 mo. Mean plasma cholesterol at ages 6 and 12 mo in the remaining five infants (158*21; 163*30 mg/lOO ml) was considerably lower than in familial Type II, but showed the same pattern of response to diet. Males vs. Females: In view of the reported sex-related difference in plasma cholesterol in infancy 8 effects of diet on plasma cholesterol were compared in males and females (Table 6). No significant differences in cord blood cholesterol, or plasma cholesterol at ages 6 and 12 mo were perceived. DISCUSSION
Although the exact type of diet was not prescribed for these infants studied, selection of infants taking either cow or breast milk or vegetable oil-skim milk formula allowed a comparison of two dietary groups: moderate/moderate and low/moderate dietary groups for normal and hypercholesterolemic infants. In both normal and hypercholesterolemic infants, the gradual increase in dietary cholesterol and saturated fatty acids in the first yr of life is accompanied by an increase from mean cord blood choIesterol levels of 64 mg/ Table 5. Cholesterol
Content of Strained Baby Food (mg/lOO g)
Food
Cholesterol
Food
Cholesterol
Egg Yolk Liver
352 300
Beef Ham
60 60
Vanilla Custard
38
Pudding’ Chicken
130 102
Veal Lamb
80 60
Cottage Cheese Peast
27
‘With eggs. tCooked with butter.
Food
Cholesterol
16
1190
GLUECK
ET AL.
100 ml (for normals) or 119-129 mg/lOO ml (for hypercholesterolemic infants) to levels of 128-129 mg/lOO ml for normals (at age 12 mo) and 197-200 mg/ 100 ml for hypercholesterolemic infants (at age 12 mo). This gradual increase in plasma cholesterol in normal and hypercholesterolemic neonates did not, under the conditions of this study, appear to be affected by early exposure to low or moderate cholesterol intake followed by ingestion of substantially increased cholesterol in the second 6 mo of life. For infants with proven neonatal and familial Type II, low cholesterol intake in early neonatal periods did not impair subsequent response to increased dietary cholesterol. At 12 mo of age (and after 6 mo of moderate cholesterol intake), familial Type II infants had the highest plasma cholesterol values (mean 225 mg/lOO ml), Comparable results (but on a much shorter study period) were obtained by Lindquist and Malmcrona,5 who gave a corn oil base formula to five infants for 8 days, followed by 2 wk of cream or breast milk. At followup, total lipids and plasma cholesterol in infants progressing from corn oil to cream-breast milk could not be differentiated from those of children on breast milk throughout. These findings in infants differ appreciabIy from studies in rat&O and pigs9 where it was postulated that cholesterol in the diet of the suckling controls development of enzymes concerned with cholesterol homeostasis in the adult. Kubat et al. weaned rats at 18 and 30 days, then fed a “high fat diet” until 10 mo of age, followed by 2% mo of cholesterol-rich diet. Although there were no group differences in serum cholesterol at 10 mo, serum cholesterol was higher at 12% mo in the early weaned group.lO Reiser fed a small number of pigs low cholesterol (1.5 mg/dl) or moderate cholesterol (114 mg/dl) milk, followed by normal pig feed and serial increases in addi-
Table 6
Neonatal Hypercholesterolemia Comparison of cord blood cholesterol, plasma cholesterol at 6 and 12 mo, males vs. females (moderate/moderate diet group) Males (N=5)
Cord blood cholesterol (mg/lOO ml) Plasma cholesterol, age 6 mo Plasma cholesterol, age 12 mo
111.6 2 9.3 166.3 k 31 165 k-l.6
Females (N=6)
Significance
125.2 -c 15.9 206.6 & 31 223 ‘- 60
Normocholesterolemic Neonates Comparison of cord blood cholesterol, plasma cholesterol at 12 mo, males vs. females (moderate/moderate diet group) Females Males (N=4)
Cord blood cholesterol (mg/lOO ml) Plasma cholesterol, age 12 mo
63 122
-t 8.8 2 36.2
Significance
(N-4)
65 133
k 6.6 s 16.5
Normorcholesterolemic Neonates Comparison of cord blood cholesterol, plasma cholesterol 12 mo, males vs. females (low/moderate diet group)
Cord blood cholesterol (mg/lOO ml) Plasma cholesterol, age 12 mo
NS NS NS
NS NS
at
Males (N=5)
Females (N=8)
55.8 -1- 15.9 140 r+ 25.7
69.1 -c 11.6 122.5 & 25.9
Significance
NS NS
DIETARY
CHOLESTEROL
1191
IN INFANCY
tional exogenous cholesterol. At 17 mo, cholesterol levels were higher in female pigs originally maintained on low cholesterol milk.9 Some of the apparent dichotomy between human and animal data may lie in the timing and duration of both the “suckling period” and the life span. The age of resampling is obviously critical; Kubat found no differences between dietary groups at age IO mo, with appreciable differences at 12% mo. Extended followup on ad lib. diets with 600 mg cholesterol/day (or more) will be necessary in our infant group to ascertain if the effects of “early low cholesterol diets” are not expressed.much later in the child’s life, rather than 5-6 mo after the change in diet. Other reservations as to the role of the “low cholesterol, polyunsaturaterich diet” in normal or hypercholesterolemic children have centered about the findings that in several animal speciesz3 increased levels of unsaturated fat lead to increased requirement for Vitamin E. Moreover, Hassan et al. have described a syndrome involving edema, papular erythema, an elevated platelet count, and anemia in a small series of premature infants receiving formula relatively rich in polyunsaturated fatty acids.2” Dayton et al. have also described in an elderly population lowered plasma Vitamin E and increased red cell hemolysis during a low cholesterol diet, rich in polyunsaturates.25 Athough comparable changes have not been reported in term infants on less radical dietary formulae than that used by Hassan et a1.,“4 approximately 60% of infants on low cholesterol, polyunsaturate-rich formulae in our studies have received additional Vitamin E supplementation, and none (with or without Vitamin E supplementation) developed the lesions or syndrome described above. Against the potential and for the most part still theoretical problems encountered with early low cholesterol polyunsaturate-rich feeding must be weighed the potential benefit of amelioration of hypercholesterolemia in neonatal familial Type II,ls7 and the much lower cholesterol levels in the normocholesterolemic child’-6*8 on low cholesterol diets. It has been suggested that dietary intervention in the infant with familial type II be started at 1 yr of age 26,27 although our experience suggests that modest intervention in the for6 of low cholesterol, polyunsaturate-rich milks can profitably be started even earlier.lf7 For the normal child, prudent restriction in dietary cholesterol intake might potentially be useful in a general reduction in plasma cholesterol levels. Primary prevention of atherosclerosis in normal and hypercholesterolemic children may potentially start in childhood. Long-term (generation) studies in normal and hypercholesterolemic children will need to be carried out to determine the efficacy (or lack of efficacy) of long-term cholesterol lowering regimens in atherosclerosis prevention. Under the restricted, short-term conditions of this study, very early low cholesterol intake does not appear to adversely affect subsequent ability to maintain “normal cholesterol homeostasis” when cholesterol intake increases in later infancy. REFERENCES 1. Glueck, C. J., and Tsang, R. C.: Neonatal Familial Type II Hyperlipoproteinemia: Effects of diet on plasma cholesterol
in the first year of life. Amer. 25 :224,
1972.
J. Clin. Nutr.
1192 2. Woodruff, C. W., Bailey, M. C., Davis, J. T., Rogers, N., and Conglio, J. G.: Serum lipids in breast-fed infants and in infants fed evaporated milk. Amer. J. Clin. Nutr.
14:83,1964. 3.Lowe, C. J., Mosovich,
proteins
in human
serum.
GLUECK
ET AL.
J. Clin.
Invest.
34:1345,1955. 15. Kessler, G., and Lederer, metric Skeggs,
measurement L. T. Jr.
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