Giovanna Marchini

Antimicrobial polypeptides of human vernix caseosa and amniotic fluid: implications for newborn innate defense.

Antimicrobial peptides/proteins are widespread in nature and play a critical role in host defense. To investigate whether these components contribute to surface protection of newborns at birth, we have characterized antimicrobial polypeptides in vernix caseosa (vernix) and amniotic fluid (AF). Concentrated peptide/protein extracts were obtained from 11 samples of vernix and six samples of AF and analyzed for antimicrobial activity using an inhibition zone assay. Proteins/peptides in all vernix extracts exhibited strong antibacterial activity against Bacillus megaterium (strain Bm11), in addition to antifungal activity against Candida albicans, whereas AF-derived proteins/peptides showed only the former activity. Fractions obtained after separation by reverse-phase HPLC exhibited antibacterial activity, with the most pronounced activity in a fraction containing α-defensins (HNP1-3). The presence of HNP1-3 was proved by dot blot analysis and confirmed by mass spectrometry. Lysozyme and ubiquitin were identified by sequence analysis in two fractions with antibacterial activity. Fractions of vernix and AF were also positive for LL-37 with dot blot and Western blot analyses, and one fraction apparently contained an extended form of LL-37. Interestingly, psoriasin, a calcium-binding protein that is upregulated in psoriatic skin and was found recently to exhibit antimicrobial activity, was characterized in the vernix extract. The presence of all of these antimicrobial polypeptides in vernix suggests that they are important for surface defense and may have an active biologic role against microbial invasion at birth.

The newborn infant is protected by an innate antimicrobial barrier: peptide antibiotics are present in the skin and vernix caseosa.

Summary Background  Peptide antibiotics are part of the surface defences against microbial intruders. However, the presence and significance of these innate immune effectors in the skin barrier of the newborn infant have not yet been appreciated. Erythema toxicum neonatorum is an inflammatory skin reaction of unknown aetiology and significance, commonly present in the healthy newborn infant.

Release of GI Hormones in Mother and Infant by Sensory Stimulation

ABSTRACT. It is well established that sensory stimulation is of great importance for the growth of and for the physiological and psychological development of infants. Supplementary sensory stimulation such as non‐nutritive sucking and tactile stimulation has been shown to increase the growth rate and the maturation of premature infants. In human neonates non‐nutritive sucking has a vagally mediated influence on the levels of some gastrointestinal hormones. In animal experiments afferent electrical stimulations of the sciatic nerves at low intensity leads to an activation of the vagal nerves and to a consequent release of vagally controlled gastrointestinal hormones such as gastrin and cholecystokinin. We therefore assume that both nonnutritive sucking and tactile stimulation trigger the activity of sensory nerves which leads to a release of vagally regulated gut hormones. Since gut hormones stimulate gastrointestinal motor and secretory activity and the growth of the gastrointestinal tract, and enhance the glucose‐induced insulin release, they may contribute to the beneficial effects on maturation and growth caused by sensory stimulation. In the breast‐feeding situation, the sucking of the child elicits similar reflexes in the mother leading to an activation of the maternal gut endocrine system and a consequent increase in energy uptake. These data indicate that many types of neurogenic reflexes induced in mother‐infant interactions are of importance for the energy economy of both mother and child.

Human cathelicidin peptide LL37 inhibits both attachment capability and biofilm formation of Staphylococcus epidermidis.

Aims:  The aim of this work was to investigate the possible effect of human cathelicidin antimicrobial peptide LL37 on biofilm formation of Staphylococcus epidermidis, a major causative agent of indwelling device‐related infections.

Erythema Toxicum Neonatorum Is an Innate Immune Response to Commensal Microbes Penetrated into the Skin of the Newborn Infant

Erythema toxicum neonatorum is a common rash of unknown etiology affecting healthy newborn infants. In this study, we postulated that the rash reflects a response to microbial colonization of the skin at birth, and that the hair follicle constitutes an “easily opened door” for microbes into the skin of the newborn. We collected microbial cultures from the skin of 69 healthy, 1-d-old infants with and without erythema toxicum to identify the colonizing flora and correlate culture results with clinical findings. We also analyzed biopsies from lesions of erythema toxicum with scanning and transmission electron microscopy in the search for microbes. Finally, each infants body temperature was measured as a sign of acute phase response. We found that 84% of 1-d-old healthy infants, with and without erythema toxicum were colonized with coagulase-negative staphylococci. In all lesions of erythema toxicum, TEM identified cocci-like bacteria localized in the hair follicle epithelium and into recruited immune cells surrounding the hair follicle; morphology and dimension supported their identification as belonging to the genus Staphylococcus. SEM revealed 10 times more hair structures per skin surface unit in newborns compared with adults. Infants with erythema toxicum also had higher body temperature. In erythema toxicum, commensal microbes gain entry into the skin tissue, most probably through the hair canal. This triggers the local immune system and a systemic acute phase response, including an increase in body temperature. We speculate that early microbial exposure to the newborn may be important for the maturation of the immune system.

Nonnutritive sucking in tube-fed preterm infants: effects on gastric motility and gastric contents of somatostatin

The present study investigated the way that sucking of a pacifier influences gastric secretory and motor functions in connection with tube feeding. Experiments were performed on eight preterm infants who were tube fed twice--once with and once without sucking of a pacifier. The time for tube feeding was significantly decreased and gastric retention decreased in five of seven infants when sucking a pacifier. Maternal milk was found to contain gastrin-17, somatostatin-14, and a somatostatin-like peptide larger than somatostatin-28. Somatostatin levels were significantly reduced in connection with non-nutritive sucking. Gastrin levels were increased in six of ten experiments 2 h and/or 3 h after bolus feeding, suggesting that these peptides were not only supplied by the milk, but were also released from the gastric mucosa. The presence of gastrin and somatostatin in gastric aspirates was established by use of chromatographic methods. The results indicate that somatostatin and gastrin are released into the gastric lumen in preterm infants and that sucking of a pacifier, in connection with bolus feeding, stimulates the gastric motor functions and facilitates the digestion process, probably via activation of vagal mechanisms.

Antimicrobial components of the neonatal gut affected upon colonization

Antimicrobial peptides (AMP) produced throughout our body are important effectors in the defense barrier of innate immunity. Here, we have analyzed antimicrobial activity and polypeptide composition of meconium versus neonatal feces to address the development of antimicrobial defense of the neonatal gut. Extracts of meconium exhibited antimicrobial activity against Bacillus megaterium, Escherichia coli, and group B streptococci (GBS) but not against the yeast Candida albicans. Extracts of neonatal feces were found to possess low activity against E. coli, GBS, and C. albicans. However, the anti-B. megaterium activity was higher in the fecal extracts than in meconium. All activities were reduced or abolished when salt was added to the antimicrobial assay. The AMP cathelicidin LL-37, α-defensin HNP-1-2, α-defensin HD 5, and lysozyme were identified in both meconium and fecal extracts. In addition, HNP-3 and a fragment of azurocidin were found in meconium, whereas the holoprotein azurocidin was detected in feces. In meconium, histones H2A and H4 were isolated and identified by their antimicrobial activity. Notably, LL-37 and lysozyme were found at significantly higher levels in feces than in meconium. Our findings reveal that meconium and feces contain AMP, acting in the defense of the neonatal gut, and may be implicated in the control of the initial colonization.

Plasma cholecystokinin concentrations after breast feeding in healthy 4 day old infants.

The aim of the present study was to characterise plasma concentrations of cholecystokinin (CCK) after breast feeding in newborn infants. Fifty eight healthy full term exclusively breast fed infants were investigated at 4 (1) (2-6) days of age. Each infant contributed one blood sample collected just before, immediately after, or 10, 30, and 60 minutes after breast feeding. Plasma concentrations of CCK were measured with a technique consisting of high pressure liquid chromatography separation of gastrins and CCKs and consequent analysis with radioimmunoassay. Mean (SD) preprandial plasma concentrations of CCK (CCK8+CCK-33,39) were 68 (17) pmol/l. A significant increase was seen immediately after breast feeding, which was followed by a decline at 10 minutes and a secondary rise was seen at 30 and 60 minutes. The first peak is likely to be due to a suckling related activation of the vagal nerve and the second to a stimulatory effect of food on CCK-producing cells. An inverse relationship between basal concentrations of CCK and age of the infant was found. In rats peripheral injections of CCK reduce food intake and cause postprandial sedation and sleepiness via activation of an afferent vagal mechanism. CCK release in response to breast feeding may therefore in addition to exerting stimulatory effects on digestion and metabolism contribute to relaxation and sleepiness seen after breast feeding. The high CCK concentrations seen in younger infants may help the infant to remain satiated and calm despite receiving very little food during the first days of life.

Erythema toxicum neonatorum: an immunohistochemical analysis.

Abstract: Erythema toxicum neonatorum is a benign rash of unknown etiology, present to various degrees in most term newborns and characterized by an accumulation of eosinophils in dermal lesions. The recruitment of leukocytes to tissues implicates the involvement of adhesion molecules, cytokines, and chemokines. We therefore performed immunohistochemistry on punch biopsy specimens from cutaneous lesions of ten 1‐day‐old infants with erythema toxicum using specific monoclonal antibodies directed against a variety of adhesion molecules, cytokines, chemokines, and cell type‐specific membrane markers. Biopsy specimens of noninflamed skin from four matched newborns and four adults served as controls. The immunohistologic features of erythema toxicum in all 10 infants included a strong staining of the adhesion molecule E‐selectin in the vessel wall and the presence of numerous inflammatory cells that were identified as dendritic cells (CD1a, CD83, HLA‐DR, CD40, and ICAM‐1 positive), eosinophils (EG2 positive), neutrophils (CD15 positive), macrophages (CD14, CD68, and Mac387 positive), and E‐selectin‐expressing cells. Furthermore, the lesions showed a high incidence of the proinflammatory cytokines interleukin (IL)‐1α and IL‐1β and of the chemokines IL‐8 and eotaxin. This immunologic activity was reduced or absent in noninflamed skin from newborn controls and adults. We conclude that there is an accumulation and activation of immune cells in the lesions of erythema toxicum, also present in noninflamed skin of 1‐day‐old infants, but to a lower level. The physiologic significance of the rash remains to be elucidated.

Fetal and maternal plasma levels of gastrin, somatostatin and oxytocin after vaginal delivery and elective cesarean section

We measured the cord levels of gastrin, somatostatin and oxytocin with radioimmunoassay in plasma collected from the umbilical artery after vaginal delivery and after elective cesarean section. Maternal venous samples after the two labour modalities were also assayed for the same hormones. Fetal gastrin, somatostatin and oxytocin levels were significantly higher after vaginal delivery than after elective cesarean section. Independently of labour type, the fetal gastrin and somatostatin levels were always higher than the maternal levels. We suggest that the observed high levels of gastrin, somatostatin and oxytocin could be due to a stress-related stimulation of the oxytocin- as well as of the gastric gastrin- and somatostatin-producing cells, occurring particularly during vaginal delivery. The significant inverse correlation found between fetal pH and the recorded hormone levels is consistent with this hypothesis.