Daniela Fanni

Marked interindividual variability in renal maturation of preterm infants: lessons from autopsy

The kidney of low birthweight preterm infants is characterized by a reduced number of mature nephrons at birth. The aim of the present study was to determine whether, in preterms, active glomerulogenesis occurs in the postnatal period and whether it may compensate the reduced number of nephrons developed during the intrauterine life. Kidney samples were obtained at autopsy from 8 human fetuses, 12 premature infants, and 3 term newborns. Glomerulogenesis, as measured by radial glomerular count (RGC), was markedly decreased in all preterm infants as compared with term newborns. A marked interindividual variability was detected in the level of glomerulogenesis, which, in the vast majority of cases, did neither correlate with the gestational age at birth nor with birthweight. Active glomerulogenesis, as demonstrated by the presence of S-shaped bodies in the subcapsular region, was present in all preterm infants in the perinatal period, but it ceased in a preterm surviving for 3 months. Our data show that active glomerulogenesis continues even after birth for a short period, although it is not able to compensate a marked oligonephronia at birth. As a consequence, the incomplete nephrogenesis typical of all extremely low birthweight preterm infants possibly results in a persistent oligonephronia which should likelihood represent a major risk factors of progressive renal disease in adulthood.

Morphogenesis and molecular mechanisms involved in human kidney development

The development of the human kidney is a complex process that requires interactions between epithelial and mesenchymal cells, eventually leading to the coordinated growth and differentiation of multiple highly specialized stromal, vascular, and epithelial cell types. The application of molecular biology and immunocytochemistry to the study of cell types involved in renal morphogenesis is leading to a better understanding of nephrogenesis, which requires a fine balance of many factors that can be disturbed by various prenatal events in humans. The aim of this paper is to review human kidney organogenesis, with particular emphasis on the sequence of morphological events, on the immunohistochemical peculiarities of nephron progenitor populations and on the molecular pathways regulating the process of mesenchymal to epithelial transition. Kidney development can be subdivided into five steps: (i) the primary ureteric bud (UB); (ii) the cap mesenchyme; (iii) the mesenchymal–epithelial transition; (iv) glomerulogenesis and tubulogenesis; (v) the interstitial cells. Complex correlations between morphological and molecular events from the origin of the UB and its branching to the metanephric mesenchyme, ending with the maturation of nephrons, have been reported in different animals, including mammals. Marked differences, observed among different species in the origin and the duration of nephrogenesis, suggest that morphological and molecular events may be different in different animal species and mammals. Further studies must be carried out in humans to verify at the morphological, immunohistochemical, and molecular levels if the outcome in humans parallels that previously described in other species. J. Cell. Physiol. 227: 1257–1268, 2012.

Thymosin β4 and β10 Levels in Pre-Term Newborn Oral Cavity and Foetal Salivary Glands Evidence a Switch of Secretion during Foetal Development

Background Thymosin β4, its sulfoxide, and thymosin β10 were detected in whole saliva of human pre-term newborns by reversed-phase high performance chromatography coupled to electrospray ion-trap mass spectrometry. Methodology/Principal Findings Despite high inter-individual variability, concentration of β-thymosins increases with an inversely proportional trend to postmenstrual age (PMA: gestational age plus chronological age after birth) reaching a value more than twenty times higher than in adult whole saliva at 190 days (27 weeks) of PMA (thymosin β4 concentration: more than 2.0 µmol/L versus 0.1 µmol/L). On the other hand, the ratio between thymosin β4 and thymosin β10 exhibits a constant value of about 4 along all the range of PMA (190–550 days of PMA) examined. In order to investigate thymosin β4 origin and to better establish the trend of its production as a function of gestational age (GA), immunohistochemical analysis of major and minor salivary glands of different pre-term fetuses were carried out, starting from 84 days (12 weeks) of gestational age. Reactive granules were seen in all glands with a maximum of expression around 140–150 days of GA, even though with high inter- and intra-individual variability. In infants and adults reactive granules in acinar cells were not observed, but just a diffuse cytoplasmatic staining in ductal cells. Significance This study outlines for the first time that salivary glands during foetal life express and secrete peptides such as β-thymosins probably involved in the development of the oral cavity and its annexes. The secretion increases from about 12 weeks till to about 21 weeks of GA, subsequently it decreases, almost disappearing in the period of expected date of delivery, when the gland switches towards the secretion of adult specific salivary peptides. The switch observed may be an example of further secretion switches involving other exocrine and endocrine glands during foetal development.

Factors influencing the development of a personal tailored microbiota in the neonate, with particular emphasis on antibiotic therapy.

Abstract In recent years, it has been clearly evidenced that most cells in a human being are not human: they are microbial, represented by more than 1000 microbial species. The vast majority of microbial species give rise to symbiotic host-bacterial interactions that are fundamental for human health. The complex of these microbial communities has been defined as microbiota or microbiome. These bacterial communities, forged over millennia of co-evolution with humans, are at the basis of a partnership with the developing human newborn, which is based on reciprocal molecular exchanges and cross-talking. Recent data on the role of the human microbiota in newborns and children clearly indicate that microbes have a potential importance to pediatrics, contributing to host nutrition, developmental regulation of intestinal angiogenesis, protection from pathogens, and development of the immune system. This review is aimed at reporting the most recent data on the knowledge of microbiota origin and development in the human newborn, and on the multiple factors influencing development and maturation of our microbiota, including the use and abuse of antibiotic therapies.

Overlapping between CYP3A4 and CYP3A7 expression in the fetal human liver during development

Abstract Objective: The cytochrome P450 (CYP450) superfamily is implicated in important life processes, including metabolism of many molecules. CYP3A account for the largest portion of CYP450 proteins in human, including CYP3A4, CYP3A5 and CYP3A7. The purpose of this study was to investigate the immunohistochemical expression of CYP3A4 and CYP3A7 in human liver at different post-conceptional (PC) ages. Methods: Human liver samples from 30 fetuses and newborns were, clustered according with the PC age, routinely processed for immunohistochemical analysis of CYP3A4 and CYP3A7. Results: CYP3A4 was positive in all but two cases, CYP3A7 was positive in all but one case, which was negative also for CYP3A4. Conclusions: Our data on immunohistochemical detection of CYP3A4 and CYP3A7 during development show that CYP3A4 expression is not restricted to the post-natal age, being the immunostaining for both CYP3A4 and CYP3A7 identical after 25 weeks of PC age, thus the relationship between these CYP450 isoforms should be considered much more complex than previous thought. A high interindividual variability was observed among subjects at all gestational age. The variable CYP3A expression suggests the existence of a marked interindividual variability in drug metabolism during the intrauterine life and in perinatal period.

“Physiological” renal regenerating medicine in VLBW preterm infants: could a dream come true?

An emerging hypothesis from the recent literature explain how specific adverse factors related with growth retardation as well as of low birth weight (LBW) might influence renal development during fetal life and then the insurgence of hypertension and renal disease in adulthood. In this article, after introducing a brief overview of human nephrogenesis, the most important factors influencing nephron number at birth will be reviewed, focusing on the “in utero” experiences that lead to an increased risk of developing hypertension and/or kidney disease in adult. Since nephrogenesis in preterm human newborns does not stop at birth, but it continues for 4–6 weeks postnatally, a better knowledge of the mechanisms able to accelerate nephrogenesis in the perinatal period, could represent a powerful tool in the hands of neonatologists. We suggest to define this approach to a possible therapy of a deficient nephrogenesis at birth “physiological renal regenerating medicine”. Our goal in preterm infants, especially VLBW, could be to prolong the nephrogenesis not only for 6 weeks after birth but until 36 weeks of post conceptual age, allowing newborn kidneys to restore their nephron endowment, escaping susceptibility to hypertension and to renal disease later in life.

Expression of WT1 during normal human kidney development

Wilms Tumor 1 (WT1) is a zinc finger protein, expressed by human podocytes in the adult kidney, which plays a relevant role in different phases of nephrogenesis in experimental animals. Since no data are available for specific role in the human fetal kidney, this study aimed at investigating the expression of WT1 during the different phases of nephrogenesis. To this end, the expression of WT1 was evaluated in the kidneys, from four human fetuses and two newborns. WT1 immunoreactivity was detected in all the examined kidneys, but not in the kidneys of the newborn at term. Immunostaining for WT1 was observed in podocytes of the glomeruli and in the subcapsular regions, in areas of active glomerulogenesis. The extent and the intensity of immunoreactivity for WT1 changed from one case to the next according to the different gestational age. This study confirms in human kidney the relevant role played by WT1 during nephrogenesis. Its expression pattern suggests a main role in the regulation of the process of Mesenchimal-Epithelial-Transition and in the development and maturation of podocytes. Further studies are needed to verify the correlation between the expression pattern of WT1 and that of other genes products involved in nephrogenesis, in order to better understand their relationship at protein level.

Thymosin β-4 in colorectal cancer is localized predominantly at the invasion front in tumor cells undergoing epithelial mesenchymal transition

Objective Thymosin β-4 (Tβ4) is a ubiquitous peptide that plays pivotal roles in the cytoskeletal system and in cell differentiation during embryogenesis. Recently, a role for Tβ4 has been proposed in experimental and human carcinogenesis. This study was aimed at evaluating the correlation between Tβ4 immunoractivity and colorectal cancer, with particular attemption to tumor cells undergoing epithelial-mesenchymal transition. Methods and Results 86 intestinal biopsies were retrospectively analyzed including 76 colorectal adenocarcinomas with evident features of epithelial-mesenchymal transition, and 10 samples of normal colorectal mucosa. Paraffin sections were immunostained for Tβ4 and for E-cadherin. Total RNA was isolated from frozen specimens obtained, at surgery, from the normal colon mucosa, the deeper regions and the superficial tumor regions in four cases of colon cancer. Tβ4 immunoreactivity was detected in the vast majority (59/76) of colon carcinomas, showing a patchy distribution, with well differentiated areas significantly more reactive than the less differentiated tumor zones. We also noted a zonal pattern in the majority of tumors, characterized by a progressive increase in immunostaining for Tβ4 from the superficial toward the deepest tumor regions. The strongest expression for Tβ4 was frequently detected in invading tumor cells with features of epithelial-mesenchymal transition. The increase in reactivity for Tβ4 matched with a progressive decrease in E-cadherin expression in invading cancer cells. At mRNA level, the differences in Tβ4 expression between the surrounding colon mucosa and the tumors samples were not significant. Conclusions Our data show that Tβ4 is expressed in the majority of colon cancers, with preferential immunoreactivity in deep tumor regions. The preferential expression of the peptide and the increase in intensity of the immunostaining at the invasion front suggests a possible link between the peptide and the process of epithelial mesenchymal transition, suggesting a role for Tβ4 in colorectal cancer invasion and metastasis.

The pine-cone body: an intermediate structure between the cap mesenchyme and the renal vesicle in the developing nod mouse kidney revealed by an ultrastructural study.

Nephrogenesis is mainly characterized by the interaction of two distinct renal constituents, the ureteric bud and the metanephric mesenchyme. In this paper we describe by means of light and electron microscopic techniques the morphological events that take place during the early stages of cap mesenchymal formation. Samples of normal renal tissue were excised from newborn NOD mice and processed by standard light and electron microscopy techniques. In all samples examined we detected the presence of several cap mesenchymal aggregates in different stages of differentiation. They varied from small solid nodules with few ovoid cells to bigger pine-cone-like aggregates, characterized by a peculiar distribution and morphology of their cellular constituents. Our data highlight, for the first time, the presence of a specific cap mesenchymal structure, the pine-cone body and show, at ultrastructural level, how each cap aggregate epithelializes proceeding in stages from a condensed mesenchymal aggregate to the renal vesicle, through the intermediate “pine-cone body” stage.

Expression pattern of thymosin beta 4 in the adult human liver

Thymosin beta-4 (Tβ4) is a member of beta-thymosins, a family of small peptides involved in polymerization of G-actin, and in many critical biological processes including apoptosis, cell migration, angiogenesis, and fibrosis. Previous studies in the newborn liver did not reveal any significant reactivity for Tβ4 during the intrauterine life. The aim of the present study was to investigate by immunohistochemistry Tβ4 expression in the adult normal liver. Thirty-five human liver samples, including 11 needle liver biopsies and 24 liver specimens obtained at autopsy, in which no pathological change was detected at the histological examination, were immunostained utilizing an anti-Tβ4 commercial antibody. Tβ4 was detected in the hepatocytes of all adult normal livers examined. A zonation of Tβ4 expression was evident in the vast majority of cases. Immunostaining was preferentially detected in zone 3, while a minor degree of reactivity was detected in periportal hepatocytes (zone 1). At higher power, Tβ4-reactive granules appeared mainly localized at the biliary pole of hepatocytes. In cases with a strong immunostaining, even perinuclear areas and the sinusoidal pole of hepatocytes appeared interested by immunoreactivity for Tβ4. The current work first evidences a strong diffuse expression of Tβ4 in the adult human liver, and adds hepatocytes to the list of human cells able to synthesize large amounts of Tβ4 in adulthood. Moreover, Tβ4 should be added to the liver proteins characterized by a zonate expression pattern, in a descending gradient from the terminal vein to the periportal areas of the liver acinus. Identifying the intimate role played by this peptide intracellularly and extracellularly, in physiology and in different liver diseases, is a major challenge for future research focusing on Tβ4.