Influence of hydrocortisone acetate administered to the lactating rat on protein and lactose content in milk and serum protein, glucose and insulin levels in dams and pups

The influence of i.m. administration to the mother of hydrocortisone acetate (doses of 0.4, 0.8 or 2.0 mg/100 g body weight/day) during the first 15 days of lactation on milk protein and lactose composition and serum levels of protein, glucose and insulin in dams and pups is studied. Total serum proteins and albumin/globulin ratio in dams were unchanged by treatment. The daily injection of 0.4 or 0.8 mg/100 g body weight failed to alter serum levels of glucose or insulin in dams, whereas a dose of 2.0 mg/100 g body weight led to a rise in glucemia (from 118 +/- 3.2 to 133 +/- 5.3) which was accompanied by a sharp change in insulinemia (from 40.7 +/- 4.1 to 83.6 +/- 6.9). All three doses raised protein levels in milk. The smallest increase was recorded with 2.0 mg/100 g body weight; this dose also reduced milk lactose content. Total serum proteins in pups rose slightly but nonsignificantly, and no significant effects were noted on albumin/globulin ratio or serum glucose and insulin levels.     

Effect of dietary protein, zinc, and carbon monoxide on fetal zinc concentration in mice

Dietary protein and zinc deficiencies known to be detrimental to the developing fetus are common in pregnant women in developing countries. Everyone in modern society is at risk of exposure to carbon monoxide (CO). This study was conducted to observe the effect of dietary protein, zinc, and exposure to CO on the fetal zinc concentrations by factorial experimentation. Pregnant mice of CD-1 strain were maintained on 17% (control) or 9% (deficient) protein diets mixed with deficient, normal (control), or supplemental zinc throughout gestation. The dams in each dietary group were exposed to air (control) or 500 ppm CO in air in environmental chambers from gestation day 8 to gestational day 18. The dams were sacrificed on d 18 and fetal zinc levels were measured by atomic absorption spectrophotometry. Carbon monoxide levels used in this study had no significant effect on fetal zinc concentration in any treatment group. When both dietary protein and zinc levels were normal, the mean fetal zinc concentrations were higher than all other dietary protein/zinc combinations (15.2±6.0 and 14.2±4.1 μg Zn/g of tissue for 0 and 500 ppm CO levels). However, when dietary protein levels were deficient, supplemental zinc increased the fetal zinc concentrations significantly (12.7±3.8 and 13.1±0.3.6 μg Zn/g of tissue, in 0 and 500 ppm CO groups) as compared to zinc-deficient groups (8.7±3.0 and 10.0±3.3 μg Zn/g of tissue in 0 and 500 ppm CO groups). The results of this study may be relevant to populations that experience both marginal zinc and protein diets during gestation.     

Peripartal gamma-glutamyltranspeptidase (GGT) activity in the mammary glands of pregnant Uje: WIST rats.

On Day 21 of gestation ( = expected day of delivery), GGT activity of the mammary gland is low 4-6 h before beginning of delivery. It is significantly higher immediately (up to 3 h) before delivery or after labor. The extent of prepartal increase is individually different. The activity measured early in the morning cannot be used to predict the remaining time to beginning of spontaneous parturition.     

The food intake of rats during pregnancy and lactation

Quantities of food required by Sprague-Dawley rats during gestation and lactation and in the post-lactation period were examined. Rats allowed to eat ad libitum during pregnancy consumed quantities of food only slightly greater than the amount reported to be the average intake of pregnant Sprague-Dawley rats (20 g/day). Rats delivered their pups on day 22 or day 23 of the gestation period, but regardless of the day of delivery, the food intake of each rat decreased on day 21 of pregnancy and then decreased a second time on the day of parturition. During lactation, food consumption of the rats soon exceeded the amount reported as the average intake of lactating rats (30-35 g/day). Food intake was found to escalate from 12.2 +/- 3.1 g/rat on day 0 of lactation, the lowest intake in the study, to 94.4 +/- 23.7 g on day 21 of lactation. However, in the latter part of the lactation period, the intake represented the combined food intake of dams and pups. Eight days in the post-lactation period were required for food intake of dams to return to a level near that recorded at the beginning of pregnancy.     

The effect of prolonged hypobaric hypoxia on growth of fetal sheep

The effect of prolonged hypobaric hypoxia on fetal sheep was studied. Pregnant ewes were subjected to an atmospheric pressure of 429 torr from 30 days to 135 days gestation (long-term study). Average fetal weight for the hypoxaemic group (3.35 +/- 0.53 kg; n = 4; mean +/- SD) was significantly lower than for the controls (4.23 +/- 0.29 kg; n = 7; P less than 0.05). A short-term study was undertaken with fetuses (n = 8) which were catheterized at 110 days gestation and whose dams were subjected to hypobaric hypoxia from 120 to 141 days gestation. The mean carotid PO2 of fetuses in the hypoxic group was 12.7 +/- 0.7 torr compared to 22.7 +/- 0.7 torr for the control group (n = 9; P less than 0.001) throughout the period of treatment. Fetal arterial oxygen content fell from 6.5 +/- 1.7 to 4.9 +/- 0.4 ml/dl (P less than 0.05), but rose to control values after 7 days due to an increase in fetal haemoglobin concentration (9.6 +/- 1.1 to 13.0 +/- 1.9 g/dl, P less than 0.001) and packed cell volume (33 +/- 3 to 45 +/- 4%, P less than 0.001). In the hypoxaemic fetuses, pH fell initially from 7.34 +/- 0.02 to 7.28 +/- 0.03 (P less than 0.05) and then recovered to 7.32 +/- 0.03 within 24 h. Mean fetal weight of the short-term hypoxic group was 3.46 +/- 0.72 kg compared to 4.15 +/- 0.51 for the control group (P less than 0.05). Both long- and short-term hypoxia produced a similar reduction in fetal body weight. The adrenal glands were significantly heavier in the hypoxic fetuses than in controls. Placental weight was not effected by hypoxia, but exposure from 30 days gestation reduced the average size of cotyledons (P less than 0.05). It is concluded that the fetal sheep increases its ability to acquire and transport oxygen in response to chronic hypoxia, but this compensation is not sufficient to prevent growth retardation or changes to the pattern of tissue growth.     

Gastrin releasing peptide-like immunoreactive substance in rat mammary glands during pregnancy

The concentration of gastrin releasing peptide-like immunoreactive substance (GRP-IS) was measured by enzyme immunoassay (EIA) in rat mammary glands during pregnancy and after delivery. The GRP-IS concentration was high in the middle stage of pregnancy (10-14 days of gestation) and decreased in late pregnancy to reach a plateau range. By using HPLC, it was shown that GRP(20-29) and GRP(16-29) were mainly present in rat mammary glands. Immunohistochemical study revealed that epithelial cells of rat mammary glands were stained intensely with antiserum GP-6201. These results suggest that GRP-IS is produced and secreted in epithelial cells of mammary glands and takes part in the proliferation, differentiation and hypertrophy of mammary glands.     

Glucocorticoid effects on zinc transport into colostrum and milk of lactating cows

Colostrum Zn concentrations were measured in eight randomly selected Holstein dairy cows. Overall mean Zn concentrations were highest within 12 h postpartum (257 +/- 14 microM, mean +/- SEM), fell to 141 +/- 8 microM by 24 h, and then declined at a linear rate of 30 microM/d during the following 48 h. Zn concentrations at 3 d (82 +/- 5 microM) were not different from 150-d milk samples (72 +/- microM). In a second experiment, 32 early-gestation cows were blocked by stage of lactation into four groups in a randomized block design and injected with 0, 15, 30, or 45 mg of dexamethasone. Milk and blood samples were collected at 0, 12, and 24 h after injection and analyzed for Zn, and for fat, protein, and lactose in milk. Cows administered 0 and 15 mg of dexamethasone showed no difference in milk Zn concentrations compared to pretreatment measurements; however, milk Zn concentrations in cows administered 30- and 45-mg doses increased significantly. Plasma cortisol decreased in the dexamethasone-treated cows. Plasma Zn and milk fat, protein, and lactose did not change. These data indicate that glucocorticoids can mediate Zn uptake and transport by the mammary glands of lactating cows and suggest that the high Zn concentration in colostrum could be a result of the preparturient surge of cortisol.     

High-protein diet during gestation and lactation affects mammary gland mRNA abundance, milk composition and pre-weaning litter growth in mice

We evaluated the effect of a high-protein diet (HP) on pregnancy, lactational and rearing success in mice. At the time of mating, females were randomly assigned to isoenergetic diets with HP (40% w/w) or control protein levels (C; 20%). After parturition, half of the dams were fed the other diet throughout lactation resulting in four dietary groups: CC (C diet during gestation and lactation), CHP (C diet during gestation and HP diet during lactation), HPC (HP diet during gestation and C diet during lactation) and HPHP (HP diet during gestation and lactation). Maternal and offspring body mass was monitored. Measurements of maternal mammary gland (MG), kidney and abdominal fat pad masses, MG histology and MG mRNA abundance, as well as milk composition were taken at selected time points. HP diet decreased abdominal fat and increased kidney mass of lactating dams. Litter mass at birth was lower in HP than in C dams (14.8 v. 16.8 g). Dams fed an HP diet during lactation showed 5% less food intake (10.4 v. 10.9 g/day) and lower body and MG mass. On day 14 of lactation, the proportion of MG parenchyma was lower in dams fed an HP diet during gestation as compared to dams fed a C diet (64.8% v. 75.8%). Abundance of MG α-lactalbumin, β-casein, whey acidic protein, xanthine oxidoreductase mRNA at mid-lactation was decreased in all groups receiving an HP diet either during gestation and/or lactation. Milk lactose content was lower in dams fed an HP diet during lactation compared to dams fed a C diet (1.6% v. 2.0%). On days 14, 18 and 21 of lactation total litter mass was lower in litters of dams fed an HP diet during lactation, and the pups' relative kidney mass was greater than in litters suckled by dams receiving a C diet. These findings indicate that excess protein intake in reproducing mice has adverse effects on offspring early in their postnatal growth as a consequence of impaired lactational function.     

Prolactin-dependent expression of GD1 alpha ganglioside, as a component of milk fat globule, in the murine mammary glands

Lactation-associated expression of GD1 alpha ganglioside in murine mammary glands was found to be due to the increasing specific activities of Gg4Cer alpha2,3- and GM1b alpha2,6-sialyltransferases in the glands from 12th day of gestation. The gene for GM1b alpha2,6-sialyltransferase, mST6GalNAcV, which was not detected in nonpregnant glands, appeared at 12th day of gestation and increased in the following gestational and lactation periods. At 3rd day of lactation, the amounts of lipid-bound sialic acid (LSA) in the mammary glands and milk of HR-1 mice were 99.3 +/- 8.5 microg per gram of dried tissue and 2.9 microg per ml, GD1 alpha comprising 64.0% and 80.5% of the total LSA, respectively, and GD1 alpha in milk was found to be preferentially distributed in the fat globule fraction. When the mammary epithelial cells at 15th day of gestation were cultured in prolactin- and epidermal growth factor (EGF)-containing media, the synthesis of fat globules and casein, together with the enhanced synthesis of GD1 alpha, were observed in the cells in prolactin medium, indicating that synthesis of GD1 alpha occurs in association with milk production as a prolactin-dependent event. Thus, GD1 alpha ganglioside, which is characteristically distributed in the cerebellar Purkinje cells of the murine brain, is supplied to neonates through the milk of the mother.     

Oxytocin receptors in the mammary gland and reproductive tract of a marsupial, the brushtail possum (Trichosurus vulpecula).

Previous studies of marsupial lactation have shown that the milk-ejection reflex changes in sensitivity, being greater in small mammary glands sucked by small pouch young and lesser in larger glands supplying milk to larger young. The involvement of oxytocin receptors in these changes was examined in the brushtail possum Trichosurus vulpecula. Oxytocin receptors were measured in the mammary glands, uterus, and medial vaginal sacs by radioreceptor assay, using [3H]oxytocin as radioligand. In the mammary gland, a single oxytocin binding site was found with an affinity and receptor concentration of 0.81 +/- 0.41 l/nmol and 10.2 +/- 4.8 pmol/g tissue respectively (SD, 10 possums). Competitive displacement curves with related peptides and analogs showed the following order of specificity: d(CH2)5[Tyr(Me)2,Thr4,Tyr9-NH2]-vasotocin much greater than vasotocin greater than oxytocin = Arg-vasopressin greater than mesotocin greater than [Thr4,Gly7]-oxytocin = Lys-vasopressin greater than [deamino-Pen1, O-methyl-Tyr2, Arg8]-vasopressin greater than isotocin much greater than [d(CH2)5, D-Phe2, Ile4, Ala9-NH2]-AVP. [3H]Oxytocin did not bind to vasopressin receptors in the thoracic aorta. The concentration of oxytocin receptors was very high in small mammary glands (18.6 pmol/g tissue in a 2-g gland) and decreased logarithmically as the size of the mammary gland increased. It is suggested that the changes in the sensitivity of milk ejection to oxytocin is related to the concentration of mammary oxytocin receptors. The presence of oxytocin receptors in both uterus and median vaginal sacs extends previous observations and supports the hypothesis that in marsupial parturition, the uterus and medial vaginal sacs respond as a single functional unit to oxytocin.     

C-reactive protein in rat: in development, pregnancy and effect of sex hormones

1. In Wistar strain rats, the serum concentration of C-reactive protein (CRP) was 3.6 +/- 0.8 micrograms/ml at the birth and no apparent change was observed during 15 days after delivery. 2. Thereafter it increased about 30 times rapidly until day 30 and then rather gradually reaching the adult level of 0.4-0.8 mg/ml. 3. Before delivery, there were two peaks in the CRP level, on day 0 and day 15 of gestation. The concentrations were 0.70 +/- 0.06 and 0.77 +/- 0.10 mg/ml, respectively. 4. The CRP level decreased to 0.42 +/- 0.05 mg/ml at the delivery and increased to 0.54 +/- 0.05 mg/ml within 2 days after delivery. 5. The treatment with estradiol-17 beta resulted in the decrease of CRP (52%) in both ovariectomized and non-treated female rats. 6. The treatment with testosterone resulted in the increase of CRP in male but not in female rats. 7. However, in ovariectomized rats, testosterone elevated the serum CRP. 8. In neither ovariectomized nor intact female rats, progesterone and corticosterone showed any remarkable effect.     

Effect of prolonged glucose infusion into fetal sheep on body growth, fat deposition and gestation length

Eleven Merino sheep fetuses were supplemented with glucose by direct continuous intravenous infusion of 50% dextrose into the fetus from day 115 of gestation until spontaneous delivery. Infusion rates of 15 or 25 g/day per kg were used and equivalent volumes of saline were infused into 11 control fetuses. Infusion periods approximated 27 days in both groups. Fetal plasma glucose concentrations were significantly (P less than 0.001) elevated throughout glucose infusion and resulted in variable but consistently higher plasma insulin concentrations in the glucose than in the saline-infused fetuses. Glucose-infused fetuses were significantly heavier than controls (mean +/- SEM; 3.86 +/- 0.16 vs 3.28 +/- 0.24 kg, P less than 0.05) and body fat depots (in g/kg body wt.) were larger in glucose-infused than control fetuses (9.91 +/- 0.65 vs 6.73 +/- 0.37, P less than 0.005, for internal brown fat depots; 1.25 +/- 0.44 vs 0.27 + 0.13, P less than 0.05, for subcutaneous white adipose tissue). The results indicate that growth and lipid deposition in the sheep fetus are responsive to increased glucose supply, an effect which may be mediated through the actions of insulin. Mean gestation length was 146.60 +/- 1.45 days for controls and 144.18 +/- 1.23 days for glucose-infused animals (normal term 150 days).     

Initiation of fatty acid synthesis in rat mammary glands.

The rate of fatty acid synthesis from [6-14C]glucose in mammary tissue remained low until parturition at 22 days of gestation and increased 10-fold at 1 day post partum. Administration of progesterone on days 20 and 21 or removal of pups at parturition abolished this increase. In the latter case, administration of prolactin, corticosterone or oxytocin had no stimulatory effect; tissue from suckled glands in which the ducts had been ligated at parturition also showed no increased rate in 24 h. Foetoplacentectomy on day 18 did not stimulate fatty acid synthesis but subsequent suckling by foster pups did. Whereas lactose synthesis is initiated by withdrawal of progesterone from the circulation, a further stimulus related to removal of milk by suckling is required to initiate fatty acid synthesis.     

Dietary trans-vaccenic acid (trans11-18:1) increases concentration of cis9,transll-conjugated linoleic acid (rumenic acid) in tissues of lactating mice and suckling pups

Lactating mice were fed trans-vaccenic acid (trans 11-18:1, TVA) to assess desaturation of TVA to cis9,trans11-conjugated linoleic acid (9/11CLA). Diets contained 30 g x kg(-1) 18:2n-6 (LA) or 20 g LA plus 10 g 18:0 (SA), TVA, or a CLA mixture (MCLA). Compared with SA, feeding TVA increased 9/11CLA concentrations in blood plasma phospholipid, triglyceride, and free fatty acid fractions. However, concentrations of 9/11CLA in plasma fractions were greater when MCLA was fed compared with SA or TVA. No 9/11CLA was detected in liver of mice fed SA, and it was only 1 mg x g(-1) of total fatty acids in the carcass. In contrast, 9/11CLA content of liver (5 mg x g(-1)) and carcass (6 mg x g(-1)) of mice fed TVA was similar to liver (5 mg x g(-1)) and carcass (7 mg x g(-1)) of mice fed MCLA. Mammary tissue of SA-fed mice had no detectable 9/11 CLA, compared with 5 or 14 mg x g(-1) for TVA or MCLA-fed mice. Stearoyl-CoA desaturase activity in mammary tissue from TVA-fed dams was 14% greater compared with SA. Activity of this enzyme in liver tissue was similar among treatments. In pups nursing TVA-fed dams, 9/1 ICLA accounted for 3 mg x g(-1) in liver but no 9/11CLA was detected in the carcass. In pups nursing MCLA-fed dams, however, 9/11CLA accounted for 8 and 6 mg x g(-1) in liver and carcass. Results indicated TVA desaturation enhanced 9/11CLA in tissues and milk fat.     

Expression of Lysostaphin in Milk of Transgenic Mice Affects the Growth of Neonates

As an initial step towards enhancing mastitis resistance in dairy animals, we generated BLG-Lys transgenic mice that secrete lysostaphin, a potent antistaphylococcal protein, in their milk. In the current study, we continue our assessment of lysostaphin as a suitable antimicrobial protein for mastitis resistance and have investigated mammary gland development and function in three lines of transgenic mice. As the lines were propagated, there was a tendency for fewer BLG-Lys litters to survive to weaning (51% as compared to 90% for nontransgenic lines, p = 0.080). Nontransgenic pups fostered on dams from these three lines exhibited diminished growth rates during the first week of lactation. Rates of gain became comparable to pups on nontransgenic dams at later time points. Initial slow growth also resulted in decreased weaning weights for pups nursed by transgenic dams (15.35±0.27 g) when compared to pups delivered and nursed by nontransgenic dams (18.61 ± 0.61 g; p < 0.001), but the effect was temporary, as similar weights were attained by adulthood. Milk yield at peak lactation was not different between BLG-Lys (0.79 ± 0.33 g) and nontransgenic (0.91 ± 0.38 g; p = 0.166) dams. Histological examination of the transgenic mammary glands during gestation revealed no differences when compared to control glands; however, at early lactational stages, the BLG-Lys glands exhibited less alveolar area than control glands and a delay in lobulo-alveolar maturation. The results clearly demonstrate reduced growth of neonates on BLG-Lys dams; whether the poor pup performance can be attributed to delayed mammary development or the gland development merely reflects reduced suckling stimuli from the pups remains to be determined.Authors Abhijit Mitra and Kathleen S. Hruska contributed equally to this work     

Ontogeny of vasoactive intestinal peptide in the human fetal digestive tract

Immunocytochemistry and radioimmunoassay were used to assess the appearance time and tissue distribution of vasoactive intestinal peptide (VIP) in the digestive tract of the human fetus. By radioimmunoassay, VIP was measurable from 10 weeks of gestation. The peptide was abundantly distributed in the jejuno-ileum and colon, where the tissue peptide concentration rose from 9-14 weeks of gestation (18.4 +/- 4.4 and 22.0 +/- 5.0 pmol/g wet weight, respectively) to 15-21 weeks (83.0 +/- 21.1 and 98.6 +/- 36.4 pmol/g, respectively). Lower concentrations were recorded in pancreas from 9-14 weeks of gestation (4.3 +/- 0.8 pmol/g) to 15-21 weeks (13.9 +/- 3.7 pmol/g). The peptide concentration was 15.6 +/- 1.9 pmol/g in fundus and 25.5 +/- 3.2 pmol/g in antrum from 15 to 21 weeks of gestation. The highest concentration was recorded in duodenum from 15 to 21 weeks of gestation (118.4 +/- 40.8 pmol/g wet weight). Tissue VIP concentration and age were positively correlated in the jejuno-ileum. By immunofluorescence, immunoreactive VIP was localized in nervous fibers in the muscularis externa, in the submucosa and in the lamina propria. Scarce cell bodies were also found in the myenteric plexus. No immunofluorescent endocrine cells were observed. These results suggest: (1) the early appearance of immunoreactive VIP in gut, as early as 10 weeks of gestation; (2) the peptide, localized in nervous structures only, follows the same distribution pattern as that in adults; (3) the development of VIPergic structures is a continuous process, initiated during the 3rd month of pregnancy.     

Effects of hypergravity on mammary metabolic function: gravity acts as a continuum.

Mammary metabolic activity in pregnant rats is significantly increased in response to spaceflight. To determine whether changes in mammary metabolism are related to gravity load, we exposed pregnant rats to hypergravity and measured mammary metabolic activity. From days 11-20 of gestation (G), animals were centrifuged (20 rpm; 1.5, 1.75, or 2.0 x gravity) or were maintained at 1 G. On G20, five rats from each group were removed from the centrifuge and euthanized. The remaining dams (n = 5/treatment) were housed at 1 G until parturition. After 2 h of nursing by the pups, the postpartum dams were euthanized (G22). Glucose oxidation to CO2 and incorporation into lipids was measured. Mammary glands from dams euthanized on G20 revealed a strong negative correlation between metabolic rate and increased G load. Approximately 98% of the variation in glucose oxidation and 94% of the variation in glucose incorporation into lipids can be accounted for by differences in G load. Differences in metabolic activity disappeared in the postpartum dams. When we combined previous data from the microgravity with hypergravity environments and plotted the ratio of mammary metabolic rate vs. G load, there was a significant exponential relationship (r2 = 0.99). These data demonstrate a remarkable continuum of response across the microgravity and hypergravity environments and support the concept that gravitational load influences mammary tissue metabolism.     

Control of parturition in the sow using progesterone and prostaglandin

The effect of progesterone and prostaglandin administration on the timing of farrowing was studied in three groups of 25 sows each. Progesterone treatment (100 mg/day) on days 112, 113 and 114 of gestation (group I) significantly prolonged the gestation length to 116.4 +/- 0.4 (mean +/- s.e.) days compared to the control sows (group III; 115.5 +/- 0.2; P less than 0.05). Administration of prostaglandin (200 micrograms Cloprostanol intramuscularly) on day 115 of gestation following progesterone treatment (group II) resulted in a gestation length of 116.0 +/- 0.1 days, with the sows farrowing 25.4 +/- 1.0 h after the prostaglandin injection. 80% of the sows farrowed between 0800 and 1700 h of day 116 of gestation. Plasma progesterone levels were maintained by the exogenous progesterone during treatment. At farrowing, higher levels of progesterone were observed in groups I and II compared to controls. Prostaglandin treatment did not significantly alter withdrawal of progesterone in progesterone treated sows, suggesting that the actions of exogenous prostaglandin is primarily on the myometrium and the cervix. Hormonal treatment in late pregnancy did not have any adverse effects on piglet viability and growth rate, or subsequent reproductive performances of sows. Lactation was initiated normally, and the concentrations of lactose, protein, fat, IgG, Na+, Ca2+ and K+ in colostrum and milk were similar in all groups during the first 5 days of lactation.     

Prevention of tilorone developmental toxicity with progesterone.

The immunomodulator tilorone hydrochloride was administered (gastric intubation) once to time-pregnant Upj:TUC(SD)spf (Sprague-Dawley) rats in four experiments. In experiment 1, tilorone (250 or 500 mg/kg) was administered on day 10 of gestation. The dams were killed 4 or 72 hr after dosing. Interferon-like activity and drug levels were determined in maternal blood, spleen, and thymus, as well as in the embryos. In experiment 2, the test groups received progesterone (2 mg/kg), or tilorone (200 or 400 mg/kg), or progesterone and tilorone. The dams from each group were killed 24 or 48 hr after receiving tilorone. Experiment 3 was similar to experiment 2, except that the dams were killed on gestation day 20. In experiment 4, tilorone (400 mg/kg) was administered on gestation day 17, 18, or 19, and the dams were killed 24 hr after dosing or on gestation day 20. In all four experiments, tilorone-related maternal toxicity (regardless of whether progesterone also was administered) was observed, as characterized by marked decreases in weight gain, the occurrence of clinical signs, and in experiment 1 by decreased thymus weights, 72 hr post-dosing. Dose-related increases in the mean number of dead embryos and in serum interferon titers occurred 72 hr postdosing. In experiment 2, there was an increase in the number of dams in the 400-mg/kg (tilorone only) group with dead embryos only, 24 hr postdosing; similar results occurred in both the 200- and 400-mg/kg groups, 48 hr postdosing. However, in the groups that also received progesterone, a partial prevention of such embryolethality was evident. In experiment 3, embryotoxicity again was observed in both tilorone-treated groups, whereas several of the dams that were also given progesterone through day 19 of gestation experienced at least a partial prevention of the embryolethal effects of tilorone. In experiment 4, no fetotoxicity was observed despite the severe maternal toxicity evident.     

Quantification of surfactant pool sizes in rabbit lung during perinatal development

Methods are presented for the quantitative isolation of surfactants from fetal and newborn rabbit alveolar lavage returns and post-lavaged lung tissue homogenates. The phospholipid content of both fractions progressively increased between 27 days gestation and term (31 days). The tissue-stored fraction increased approximately 16-fold (from 0.48 +/- 0.13 to 7.83 +/- 0.86 mg/g dry lung) and the alveolar fraction more than 30-fold (from 0.08 +/- 0.02 to 2.69 +/- 0.52 mg/g dry lung). Developmental changes in phospholipid composition were also observed. Tissue-stored surfactant was prepared using differential and density gradient centrifugation. Alveolar surfactant was isolated during fetal development as a high-speed pellet following a one-step differential centrifugation. There was little change in the phospholipid content of fetal alveolar lavage supernatant (range 0.12 +/- 0.04 to 0.28 +/- 0.09 mg/g dry lung). By the first postnatal day the phospholipid content of both lavage fractions significantly increased (pellet, 7.51 +/- 1.79; supernatant, 4.01 +/- 1.36 mg/g dry lung) and both were identified as surfactant. This increase in alveolar surfactant was accompanied by an approximately twofold decrease (to 3.81 +/- 1.1 mg/g dry lung) in the tissue-stored fraction. These data provide a quantitative profile of surfactant accumulation and secretion in developing rabbit lung.