Early undernutrition attenuates the inflammatory response in adult rat offspring.

OBJECTIVE: We determined the impact of maternal food restriction during gestation and lactation or lactation alone on basal inflammation and lipopolysaccharide (LPS)-stimulated cytokine induction in adult female offspring. STUDY DESIGN: From 10 days of gestation to term, pregnant rats received either ad libitum (control) feed or were 50% food restricted (FR). Pups were either nursed by their own dams or were cross-fostered to give three groups, namely, Control (AdLib/AdLib), FR during lactation (AdLib/FR) and FR during pregnancy and lactation (FR/FR). All offspring were weaned to ad libitum rat chow. At 9 months of age, basal inflammation and LPS-stimulated cytokine responsiveness were determined in female offspring. RESULTS: The basal CRP levels were elevated in AdLib/FR and FR/FR as compared to Controls. In contrast, LPS induction of IL-1beta and IL-6 was significantly attenuated in the AdLib/FR and FR/FR offspring as compared to the Control group. CONCLUSIONS: These findings suggest that early undernutrition, particularly during prenatal and postnatal periods, affects offspring immune competence by increasing basal inflammation while reducing cytokine induction to inflammatory stimuli.

Programmed hyperphagia due to reduced anorexigenic mechanisms in intrauterine growth-restricted offspring.

Maternal food restriction during pregnancy results in intrauterine growth-restricted (IUGR) newborns with significantly decreased plasma leptin levels. When nursed by ad libitum-fed controls, IUGR offspring exhibit hyperphagia with adult obesity, marked by increased percentage body fat and plasma leptin, suggesting altered anorexigenic pathways. The authors examined leptin signaling pathways and food intake responses to 2 putative anorexic effectors (leptin and sibutramine, a serotonin reuptake inhibitor) in IUGR offspring. From 10 days to term gestation and through lactation, control pregnant rats received ad libitum food, whereas study rats were 50% food restricted. Following birth, litter size was standardized, and all offspring were nursed by control dams. At 3 weeks of age, offspring were weaned to ad libitum laboratory chow. At ages 1 day and 3 weeks, hypothalamic leptin receptor (Ob-Rb) mRNA and total STAT3 protein expression were determined. In addition, phosphorylated STAT3 was measured in 1-day-old offspring administered peripheral leptin. In prepubescent and adult offspring, anorexic effects of leptin and sibutramine were determined. At 1 day of age, IUGR pups showed increased hypothalamic Ob-Rb mRNA and total STAT3 protein expression though reduced leptin activated phosphorylated STAT3. At 3 weeks of age, IUGR offspring had decreased hypothalamic Ob-Rb mRNA expression, although with continued elevated STAT3 protein levels. The IUGR offspring demonstrated resistance to anorexigenic agents, leptin (6 weeks and 6 months), and sibutramine (8 months), as evidenced by less reduction in food intake and less body weight loss than controls. The IUGR offspring demonstrate suppressed leptin-induced STAT3 phosphorylation and impaired anorexigenic response to 2 factors in the central satiety pathway. This reduced anorexigenic function, together with normal or perhaps enhanced orexigenic function, contributes to the development of programmed obesity in IUGR rat offspring.

The timing of nutrient restriction during rat pregnancy/lactation alters metabolic syndrome phenotype.

OBJECTIVE: Modulation of growth of intrauterine growth restricted (IUGR) newborns causes either adult obesity or normalization of body weight and fat. We investigated the impact of rapid versus delayed catch-up growth of IUGR offspring on glucose and lipid profiles. STUDY DESIGN: From 10 days to term gestation and through lactation, control pregnant rats received ad libitum food, whereas study rats were 50% food restricted. Cross-fostering techniques were used to examine effects of food restriction during pregnancy and/or lactation periods. Glucose and lipid profiles were determined in offspring at ages 1 day, 3 weeks, and 9 months. RESULTS: Food restriction during pregnancy produced hypoglycemic IUGR pups. Those permitted rapid catch-up growth demonstrated adult obesity with insulin resistance (hyperglycemia/hyperinsulinemia) and hypertriglyceridemia. Conversely, IUGR exhibiting delayed catch-up growth demonstrated normal adult body weight and insulin deficiency (hyperglycemia/hypoinsulinemia) and elevated cholesterol levels as compared with controls. However, these adult offspring had higher glucose though similar insulin levels as control offspring nursed by food restricted dam. CONCLUSION: The timing and the rate of IUGR newborn catch-up growth causes markedly altered adult phenotypes. Although delayed newborn catch-up growth may be beneficial in the prevention of adult obesity, there may be significant adverse effects on pancreatic function.

A paradoxical temporal response of the PTHrP/PPARgamma signaling pathway to lipopolysaccharide in an in vitro model of the developing rat lung.

Chorioamnionitis alters lung development, resulting in a paradoxical decrease in the incidence of respiratory distress syndrome but an increase in the incidence of bronchopulmonary dysplasia (BPD). The mechanism(s) underlying this disparity in the pulmonary outcomes is not known. We hypothesized that specific alterations in alveolar epithelial-mesenchymal interactions might explain this apparent disparity in the pulmonary outcome following chorioamnionitis. We determined the effects of lipopolysaccharide (LPS) on parathyroid hormone-related protein (PTHrP)-driven epithelial-mesenchymal interactions that are essential for normal lung development and homeostasis. Lung explants from embryonic day 19.5 Sprague-Dawley rat fetuses were treated with LPS with or without a PTHrP pathway agonist, prostaglandin J(2) (PGJ(2)). LPS treatment affected the production of proinflammatory cytokines and the expression of the key markers of the epithelial-mesenchymal paracrine interactions in a time-dependent manner. At 6 h, there was a significant increase in the expression of PTHrP and the other key markers of alveolar homeostasis without any significant effect on alpha-smooth muscle actin (alphaSMA). In contrast, at 72 h, there was a significant decrease in the expression of PTHrP and the other key markers of alveolar homeostasis accompanied by a significant increase in alphaSMA expression. The cytokine and molecular changes at 72 h were completely prevented by the concomitant treatment with PGJ(2). We speculate that these data provide a likely mechanism for the acute stimulation of lung differentiation, accompanied paradoxically by BPD following chorioamnionitis, and suggest that by specifically targeting PTHrP signaling, the inflammation-induced molecular injury that is known to result in BPD can be prevented.

Maternal N-acetylcysteine suppresses fetal inflammatory cytokine responses to maternal lipopolysaccharide.

OBJECTIVE: Evidence suggests that maternal infections may induce fetal inflammatory responses. Because cytokine actions may be mediated by oxidative stress, we determined whether N-acetylcysteine, an antioxidant, can blunt fetal inflammatory responses to maternal lipopolysaccharide. STUDY DESIGN: Sprague Dawley near-term rats (n = 16) received intraperitoneal lipopolysaccharide (100 microg/kg) at 30 minutes and saline solution or N-acetylcysteine (300 mg/kg) at 150 minutes. An additional group received N-acetylcysteine before and after lipopolysaccharide administration. At 6 hours, rats were killed, and fetal and maternal blood cytokines were determined. RESULTS: After maternal lipopolysaccharide administration, fetal blood interleukin-6 markedly increased (3 +/- 2 to 1265 +/- 574 pg/mL); N-acetylcysteine that was given before or before and after lipopolysaccharide administration reduced fetal interleukin-6 response to control levels. A similar trend was observed for interleukin-1beta. No effect of N-acetylcysteine on fetal interleukin-10 levels was observed. CONCLUSION: Maternal N-acetylcysteine inhibits fetal cytokine responses to maternal lipopolysaccharide, even when given 2 hours after lipopolysaccharide injection. These results suggest that N-acetylcysteine may protect the fetus from sequelae of maternal inflammation.

Gender-specific orexigenic and anorexigenic mechanisms in rats.

Feeding dysregulation may manifest as either under-nourishment (e.g., anorexia) or excessive eating leading to obesity. Recent studies have suggested a gender-related variance in weight maintenance in response to chronic disease or obesity-related dietary regimens. However it is unclear whether these gender differences in weight management are secondary to appetite-mediated food intake or alternative mechanisms (e.g., exercise, metabolism). In this study, we explored gender-dependent feeding and hormonal responses to dietary restriction (12-h fast) or to an inflammatory stimulus (LPS, 100 microg/kg b.w.; i.p.) in rats. In response to a 12 h fast, female rats increased (p<0.05) total daily food intake above that of male rats by primarily increasing nighttime feeding by 40%, as compared to 10% in males. Consistent with the increased food intake, fasting induced a greater percent increase in female as compared to male plasma ghrelin (141 vs. 65%, p<0.001). In response to LPS, both male and female rats showed similar reductions in total daily food consumption. However LPS (6 h) induced a greater percent increase in plasma leptin in female than male rats (230 vs. 33%, p<0.01), whereas ghrelin was similarly decreased in both females and males (66 vs. 44%). These findings demonstrate sexual dimorphic responses in feeding and appetite-associated hormonal responses to fasting or LPS treatment. Our findings suggest that therapeutic interventions with ghrelin or leptin must be modified according to gender in order to optimally achieve either weight loss for obesity or weight gain/maintenance for chronic illness-associated anorexia.

N-acetyl-cysteine suppresses amniotic fluid and placenta inflammatory cytokine responses to lipopolysaccharide in rats.

OBJECTIVE: Maternal infections may induce placental, amniotic and, potentially, fetal inflammatory responses. As cytokine responses may be mediated by oxidative stress, we determined whether the antioxidant N-acetyl-cysteine (NAC), can attenuate maternally induced amniotic and placental cytokine responses to maternal infection (modeled by lipopolysaccharide [LPS]). STUDY DESIGN: Gestation day 18 pregnant rats were (1) treated with LPS (100 microg/kg, body weight; intraperitoneally) alone; (2) pretreated with NAC (300 mg/kg body weight; intraperitoneally) 30 minutes before LPS; (3) posttreated with NAC 120 minutes after LPS; or (4) treated with NAC 30 minutes before and 120 minutes after LPS. Six hours after LPS administration, maternal serum and amniotic fluid interleukin-6 (IL-6) and IL-10 levels, and placental IL-6 messenger RNA levels were determined. RESULTS: LPS increased maternal serum IL-6 (50 +/- 25 to 3444 +/- 584 pg/mL) and IL-10 (40 +/- 20 to 958 +/- 339 pg/mL) and amniotic fluid IL-6 (59 +/- 25 to 891 +/- 128 pg/mL). Pretreatment and/or posttreatment with NAC attenuated IL-6 in the maternal serum and amniotic fluid and IL-10 in the amniotic fluid. LPS also induced placental IL-6 messenger RNA that was inhibited by treatment with NAC before and after LPS. CONCLUSION: NAC inhibition of inflammatory responses may protect the fetus from potential long-term sequelae.

Permanent reduction in heart and kidney organ growth in offspring of undernourished rat dams.

OBJECTIVE: Maternal undernutrition affects fetal growth and development. We investigated whether maternal food restriction during pregnancy and/or lactation permanently alters organ growth among adult offspring. STUDY DESIGN: From 10 days to term gestation and through 21 days lactation, control pregnant rats received ad libitum food, whereas study rats were 50% food restricted. Cross-fostering techniques were used to examine the effects of food restriction during pregnancy and/or lactation periods. Organs were dissected and weighed (percentage of body weight) at ages 3 weeks and 9 months. RESULTS: Food restriction during pregnancy produced growth-restricted newborns that exhibited catch-up growth that resulted in markedly heavier adult offspring, although with relatively decreased weights of heart, kidney, lung, and brain as compared with controls. Conversely, food restriction during pregnancy/lactation or lactation alone resulted in adult offspring with similar body weights as controls, but with relatively decreased growth of heart and kidney. Males exhibited relatively smaller livers, whereas the females showed relatively smaller adrenal glands. CONCLUSION: Sex-dependent, selective, and permanent changes in relative growth of heart and kidney may increase risk of adult diseases.

Gender specificity of programmed plasma hypertonicity and hemoconcentration in adult offspring of water-restricted rat dams.

OBJECTIVE: We studied the impact of maternal water-restriction during rat pregnancy on newborn plasma composition, and determined the persistence of plasma composition alterations in adult offspring. METHODS: Maternal dams were water-restricted from 10 days of pregnancy until term (21 days) and throughout lactation to increase plasma sodium levels by approximately 6 mEq/L. At 21 days of age, offspring were weaned, and subsequently maintained on ad libitum food and water until 12 weeks of age. Daily water and food intake was monitored. Blood samples and organs were collected from 1-day- and 12-week-old offspring. Hematocrit, plasma osmolality, sodium, and arginine vasopressin (AVP) levels were analyzed. Because water-restriction led to concomitant reduction in maternal food intake (ie, dehydration anorexia), henceforth these dams and their offspring are referred to as "water-deprived/food-reduced" rats. RESULTS: Water-deprived/food-reduced dams had significantly increased plasma sodium levels, reduced food intake, and lower body weight gain during pregnancy and lactation as compared to control dams. One-day-old newborns of water-deprived/food-reduced dams weighed 17% less and had increased plasma sodium levels, osmolality, and hematocrit. At 12 weeks of age, males exhibited 11% and females 19% reduction in body weight from controls. Notably, male offspring of water-deprived/food-reduced dams showed significantly elevated plasma sodium levels, osmolality, and hematocrit. Additionally, males demonstrated reduced adrenal growth and decreased water intake. Conversely, the female offspring had similar plasma osmolality with decreased sodium levels, though a persistently elevated hematocrit. No differences were evident in plasma AVP levels. CONCLUSIONS: Maternal water deprivation/food reduction is associated with increased newborn plasma osmolality and sodium levels and long-term physiologic changes in the offspring. The gender-specificity of programmed hyperosmolality, though not hemoconcentration, implicates differing pathways/mechanisms for these phenotypic alterations. The contributions of pregnancy hypertonicity versus nutrient restriction in the mechanism for programmed offspring phenotype remain to be elucidated.

Placental and fetal membrane Nephrin and Neph1 gene expression: response to inflammation.

OBJECTIVE: Fetal and amniotic fluid (AF) proteins (eg, alpha fetoprotein [AFP]) are measurable in the maternal circulation. Elevated maternal serum AFP levels indicate a risk for fetal anomalies or for obstetrical complications that are often associated with inflammation (eg, preterm labor). However, little is known of the mechanism of protein exchange between the fetus, AF, and maternal circulation. Nephrin and Neph1 are cell membrane proteins that restrict glomerular protein filtration and which are differentially expressed with renal inflammation. We sought to investigate whether nephrin and Neph1 were expressed in placenta and fetal membranes, and whether inflammation modified the expression. METHODS: Pregnant rats at 18 days' gestation were injected with lipopolysacchride (LPS) or control saline intraperitoneally (IP) and killed at 1, 6, and 12 hours after injection. Placenta and fetal membranes were obtained and real-time polymerase chain reaction (PCR) performed for determination of nephrin and Neph1 levels. RESULTS: Nephrin and Neph1 were expressed in both placenta and fetal membranes. Following maternal LPS administration, nephrin mRNA significantly increased in the membranes (0.22 +/- 0.02 to 0.51 +/- 0.050, P <.05), while Neph1 expression significantly declined in the placenta (0.19 +/- 0.05 to 0.10 +/- 0.01, P <.05). CONCLUSION: Fetal membranes and placenta of the rat express mRNA for the protein barriers nephrin and Neph 1, suggesting a role in the regulation of protein transfer from the fetus to mother. Under basal conditions, AF AFP transfer across fetal membranes may account for maternal serum AFP levels, whereas gestational inflammatory conditions (eg, preterm labor, threatened abortion) may augment AFP transfer across the placenta.

Programmed obesity in intrauterine growth-restricted newborns: modulation by newborn nutrition.

The degree of nutrient enhancement during the newborn period may modulate programming of appetite-regulating hormones, body composition, and propensity to adult obesity in intrauterine growth-restricted (IUGR) newborns. Pregnant rats received, from day 10 to term gestation and throughout lactation, ad libitum food (AdLib) or 50% food restriction (FR) to produce IUGR newborns. AdLib vs. FR offspring were studied at day 1, and, to create two distinct groups of newborn catch-up growth (immediate, delayed) among the IUGR newborns, cross-fostering techniques were employed. The four groups of pups at 3 wk were IUGR immediate catch-up growth (FR/AdLib), IUGR delayed catch-up growth (FR/FR), control (AdLib/AdLib), and lactation FR control (AdLib/FR). From 3 wk to 9 mo, all offspring had AdLib rat chow. Maternal FR during pregnancy resulted in IUGR pups (6.0 +/- 0.3 vs. 7.1 +/- 0.3 g, P < 0.01) with decreased leptin (0.66 +/- 0.03 vs. 1.63 +/- 0.12 ng/ml, P < 0.001) and increased ghrelin (0.43 +/- 0.03 vs. 0.26 +/- 0.02 ng/ml, P < 0.001). Maternal FR during lactation (FR/FR) further impaired IUGR offspring growth at 3 wk. However, by 9 mo, these pups attained normal body weight, percent body fat, and plasma leptin levels. Conversely, IUGR offspring nursed by AdLib dams (FR/AdLib) exhibited rapid catch-up growth at 3 wk and continued accelerated growth, resulting in increased weight, percent body fat, and plasma leptin levels. Thus the degree of newborn nutrient enhancement and timing of IUGR newborn catch-up growth may determine the programming of orexigenic hormones and offspring obesity.

Central angiotensin II AT1 receptors mediate fetal swallowing and pressor responses in the near-term ovine fetus.

Swallowed volumes in the fetus are greater than adult values (per body weight) and serve to regulate amniotic fluid volume. Central ANG II stimulates swallowing, and nonspecific ANG II receptor antagonists inhibit both spontaneous and ANG II-stimulated swallowing. In the adult rat, AT1 receptors mediate both stimulated drinking and pressor activities, while the role of AT2 receptors is controversial. As fetal brain contains increased ANG II receptors compared with the adult brain, we sought to investigate the role of both AT1 and AT2 receptors in mediating fetal swallowing and pressor activities. Five pregnant ewes with singleton fetuses (130 +/- 1 days) were prepared with fetal vascular and lateral ventricle (LV) catheters and electrocorticogram and esophageal electromyogram electrodes and received three studies over 5 days. On day 1 (ANG II), following a 2-h basal period, 1 ml artificial cerebrospinal fluid (aCSF) was injected in the LV. At time 4 h, ANG II (6.4 microg) was injected in the LV, and the fetus was monitored for a final 2 h. On day 3, AT1 receptor blocker (losartan 0.5 mg) was administered at 2 h, and ANG II plus losartan was administered at 4 h. On day 5, AT2 receptor blocker (PD-123319; 0.8 mg was administered at 2 h and ANG II plus PD-123319 at 4 h. In the ANG II study, LV injection of ANG II significantly increased fetal swallowing (0.9 +/- 0.1 to 1.4 +/- 0.1 swallows/min; P < 0.05). In the losartan study, basal fetal swallowing significantly decreased in response to blockade of AT1 receptors (0.9 +/- 0.1 to 0.4 +/- 0.1 swallows/min; P < 0.05), while central injection of ANG II in the presence of AT1 receptor antagonism did not increase fetal swallowing (0.6 +/- 0.1 swallows/min). In the PD-123319 study, basal fetal swallowing did not change in response to blockade of AT2 receptor (0.9 +/- 0.1 swallows/min), while central injection of ANG II in the presence of AT2 blockade significantly increased fetal swallowing (1.5 +/- 0.1 swallows/min; P < 0.05). ANG II caused significant pressor responses in the control and PD-123319 studies but no pressor response in the presence of AT1 blockade. These data demonstrate that in the near-term ovine fetus, AT1 receptor but not AT2 receptors accessible via CSF contribute to dipsogenic and pressor responses.

Interleukin-1beta system in anorectic catabolic tumor-bearing rats.

PURPOSE OF REVIEW: The onset of cancer anorexia and the accompanying neurological symptoms and signs involve the general influence of cytokines on the brain. Using methylcholanthrene to induce tumors in Fischer 344 rats, we measured various specific components of the cytokine-induced anorectic reaction, including: (1) IL-1beta system components (ligand, signaling receptor, receptor accessory proteins, and receptor antagonist); (2) TNF-alpha; (3) TGF-beta1; and (4) IFN-gamma in the tumor tissue, the liver and the brain. RECENT FINDINGS: The data show that IL-1beta, TNF-alpha and IFN-gamma messenger RNA were detected in the tumor tissue of anorectic tumor-bearing rats. In brain regions, anorexia is associated with the upregulation of IL-1beta and its receptor mRNA. All other mRNA remained unchanged in the brain regions examined. SUMMARY: This suggests that IL-1beta and its receptor may play a significant role in this model of cancer-associated anorexia. In vivo, the characterization of cytokine components in the brain may provide data for potential pharmacological interventions to ameliorate the anorexia of disease.

Maternal LPS induces cytokines in the amniotic fluid and corticotropin releasing hormone in the fetal rat brain.

Perinatal infections are a risk factor for fetal neurological pathologies, including cerebral palsy and schizophrenia. Cytokines that are produced as part of the inflammatory response are proposed to partially mediate the neurological injury. This study investigated the effects of intraperitoneal injections of lipopolysaccharide (LPS) to pregnant rats on the production of cytokines and stress markers in the fetal environment. Gestation day 18 pregnant rats were treated with LPS (100 microg/kg body wt i.p.), and maternal serum, amniotic fluid, placenta, chorioamnion, and fetal brain were harvested at 1, 6, 12, and 24 h posttreatment to assay for LPS-induced changes in cytokine protein (ELISA) and mRNA (real-time RT-PCR) levels. We observed induction of proinflammatory cytokines interleukin (IL)-1 beta, IL-6, and tumor necrosis factor-alpha (TNF-alpha) as well as the anti-inflammatory cytokine IL-10 in the maternal serum within 6 h of LPS exposure. Similarly, proinflammatory cytokines were induced in the amniotic fluid in response to LPS; however, no significant induction of IL-10 was observed in the amniotic fluid. LPS-induced mRNA changes included upregulation of the stress-related peptide corticotropin-releasing factor in the fetal whole brain, TNF-alpha, IL-6, and IL-10 in the chorioamnion, and TNF-alpha, IL-1 beta, and IL-6 in the placenta. These findings suggest that maternal infections may lead to an unbalanced inflammatory reaction in the fetal environment that activates the fetal stress axis.

Unopposed orexic pathways in the developing fetus.

Fetal swallowing has important roles in fetal gastrointestinal development, and perhaps fetal somatic growth and maturation. Ingestive behavioral responses must develop in utero to provide for acquisition of water and food intake during the neonatal period. At birth, the rat, ovine and human fetus have developed mechanisms to acquire food via intact mechanisms of taste, suckling and swallowing. Our preliminary studies suggest that in sheep and likely in human fetuses, putative orexic-mediated ingestive responses are present near term gestation. We hypothesize that both orexic (appetite) and satiety mechanisms develop during the last third of gestation and the related neurotransmitters involved in this process are functional. The potential in utero imprinting of orexic mechanisms may influence infant, childhood and ultimately adult appetite "set-points". Thus, dysfunctional appetite, and perhaps obesity, may result from maternal environmental influences during critical stages of development.

Lipopolysaccharide (LPS)-induced dopamine cell loss in culture: roles of tumor necrosis factor-alpha, interleukin-1beta, and nitric oxide.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopamine (DA) neurons of the substantia nigra pars compacta (SNc). Although the exact mechanisms responsible for this cell loss are unclear, emerging evidence suggests the involvement of inflammatory events. In the present study, we characterized the effects of the proinflammatory bacteriotoxin lipopolysaccharide (LPS) on the number of tyrosine hydroxylase immunoreactive (THir) cells (used as an index for DA neurons) in primary mesencephalic cultures. LPS (10-80 microg/ml) selectively decreased THir cells and increased culture media levels of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) as well as nitrite (an index of nitric oxide (NO) production). Cultures exposed to both LPS and neutralizing antibodies to IL-1beta or TNF-alpha showed an attenuation of the LPS-induced THir cell loss by at least 50% in both cases. Inhibition of the inducible form of nitric oxide synthase (iNOS) by L-NIL did not affect LPS toxicity, but increased the LPS-induced levels of both TNF-alpha and IL-1beta. These findings suggest that neuroinflammatory stimuli which lead to elevations in cytokines may induce DA neuron cell loss in a NO-independent manner and contribute to PD pathogenesis.

In utero bacterial endotoxin exposure causes loss of tyrosine hydroxylase neurons in the postnatal rat midbrain.

We investigated whether in utero exposure to the Gram(-) bacteriotoxin lipopolysaccharide (LPS) induces dopamine (DA) neuron loss in rats. The proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) kills DA neurons and is elevated in the brains of patients with Parkinson's disease (PD). LPS is a potent inducer of TNF-alpha, and both are increased in the chorioamniotic environment of women who have bacterial vaginosis (BV) during pregnancy, suggesting that BV might interfere with the normal development of fetal DA neurons. Gravid female rats were injected intraperitoneally with either LPS or normal saline at embryonic day 10.5 and their pups were killed at postnatal day 21. The brains of the pups were assessed for DA and TNF-alpha levels and DA cell counts in the mesencephalon using tyrosine hydroxylase immunoreactive (THir) cells as a DA neuron marker. Prenatal LPS exposure significantly reduced striatal DA (29%) and increased DA activity (72%) as well as TNF-alpha (101%). Stereological cell counts in the mesencephalon were also significantly reduced (27%) by prenatal LPS exposure. Prenatal exposure to LPS, as might occur in humans with BV, produces a significant loss of THir cells in rats that is still present 33 days following a single injection of LPS. Since this cell loss is well past the normal phase of DA neuron apoptosis that occurs in early postnatal life, rats so exposed may have a permanent loss of DA neurons, suggesting that prenatal infections may represent risk factors for PD.

Vascular endothelial cells express isoforms of protein kinase A inhibitor.

The expression and function of the endogenous inhibitor of cAMP-dependent protein kinase (PKI) in endothelial cells are unknown. In this study, overexpression of rabbit muscle PKI gene into endothelial cells inhibited the cAMP-mediated increase and exacerbated thrombin-induced decrease in endothelial barrier function. We investigated PKI expression in human pulmonary artery (HPAECs), foreskin microvessel (HMECs), and brain microvessel endothelial cells (HBMECs). RT-PCR using specific primers for human PKI alpha, human PKI gamma, and mouse PKI beta sequences detected PKI alpha and PKI gamma mRNA in all three cell types. Sequencing and BLAST analysis indicated that forward and reverse DNA strands for PKI alpha and PKI gamma were of >96% identity with database sequences. RNase protection assays showed protection of the 542 nucleotides in HBMEC and HPAEC PKI alpha mRNA and 240 nucleotides in HBMEC, HPAEC, and HMEC PKI gamma mRNA. Western blot analysis indicated that PKI gamma protein was detected in all three cell types, whereas PKI alpha was found in HBMECs. In summary, endothelial cells from three different vascular beds express PKI alpha and PKI gamma, which may be physiologically important in endothelial barrier function.