Alice Sanna

Each niche has an actor: multiple stem cell niches in the preterm kidney

The preterm kidney cannot be simply considered as a kidney small in size: as compared to the adult kidney, the developing organ of the preterm infant is characterized by marked differences regarding the architecture and cell components. At macroscopy, fine linear demarcations indenting the renal surface characterize the fetal and preterm kidney. At microscopy, multiple major architectural changes differentiate the developing kidney from the adult one: a large capsule with a high cellularity; the branching ureteric bud, extending from the hilum towards the renal capsule; striking morphological differences among superficial (just born) and deep (more mature) glomeruli; persistence of remnants of the metanephric mesenchyme in the hylum; incomplete differentiation of developing proximal and distal tubules. At cellular level, kidneys of preterm infants are characterized by huge amounts of stem/precursor cells showing different degrees of differentiation, admixed with mature cell types. The most striking difference between the preterm and adult kidney is represented by the abundance of stem/progenitor cells in the former. Multiple stem cell niches may be identified in the preterm kidney, including the capsule, the sub-capsular nephrogenic zone, the cap mesenchyme embracing the ureteric bud tips, the cortical and medullary interstitium, and the hilar zone in proximity of the ureteric origin. The sub-capsular area represents the major stem cell niche in the prenatal kidney. It has been defined “blue strip”, due to the scarcity of cytoplasm of the undifferentiated stem/progenitors, which appear as small cells arranged in a solid pattern. All these data taken together, the morphological approach to the analysis of the preterm kidney appears completely different from that typically utilized in kidney biopsies from adult subjects. Such a different structure should be taken into account when evaluating renal function in a preterm infant in clinical practice. Moreover, a better knowledge of molecular biology of the blue strip stem/progenitor cells could be at the basis of a new “endogenous” regenerative medicine, finalized to maintain and protect the nephrogenic potential of preterm infants till the 36th week of post-conceptional age, allowing them to escape oligonephronia and chronic kidney disease later in life.

CD44 immunoreactivity in the developing human kidney: a marker of renal progenitor stem cells?

Abstract CD44 is a transmembrane adhesion glycoprotein, functioning as a hyaluronan receptor and participating in the uptake and degradation of hyaluronan. Recently, CD44 has been proposed in the adult kidney as a marker of activated glomerular parietal epithelial cells, the putative niche stem cells that, in case of damage to podocytes, might migrate inside the glomerular tuft and undergo transition to podocytes. Here, immunoreactivity for CD44 was tested in 18 human fetuses and newborns with a gestational age ranging from 11 to 39 weeks. CD44 immunoreactivity was observed in all but one developing kidneys, being localized in several renal cell types including intraglomerular, capsular, cortical and medullary interstitial cells and nerve cells. In some cases, CD44 marked scattered cells in nephrogenic subcapsular zone. Our data indicate that CD44 is involved in human nephrogenesis, probably marking a subset of progenitor/stem cells involved in early phases of kidney development and, putatively, in podocyte and/or interstitial cell differentiation.

Immunohistochemical markers of stem/progenitor cells in the developing human kidney.

The aim of this study was to better define, by immunohistochemistry, the molecular markers of renal stem/progenitor cells localized in the different niches of ten human preterm kidneys with gestational age ranging from 11 up to 25 weeks. Our data evidence the existence of multiple stem/progenitor pools in different zones of the human developing kidney that are characterized by different phenotypes: capsular stem cells were EMA (MUC1)+, MDM2+, Vimentin+ and Wnt1+; progenitors of the sub-capsular nephrogenic zone were MDM2+ and Wnt1+; cap mesenchymal cells were EMA (MUC1)+, CD15+, vimentin+, Wt1+, CD10+, Bcl2+, Wnt1+ and PAX2+; interstitial progenitor cells were Vimentin+, Wt1+ and α1Anti-tripsin+. Our data evidence the existence of multiple stem/progenitor cell pools in the fetal and neonatal human kidney. Progenitors of these different pools are characterized by a peculiar phenotype, indicating a different differentiation stage of these renal progenitors. A better knowledge of the molecular markers expressed by renal stem/progenitors might represent a relevant datum for researchers involved in renal regenerative medicine.

Decreasing podocyte number during human kidney intrauterine development.

Nephron number at birth has relevant clinical importance with implications for long-term renal health. In recent years, the podocyte depletion hypothesis has emerged as an important concept in kidney pathology. This study was aimed at verifying whether human podocyte number changes significantly during intrauterine life. To this end, 62 subjects with gestational ages ranging from 20 to 41 wk were examined. Kidney sections were stained with hematoxylin and eosin and digitally scanned at ×400 magnification. Subjects were subdivided into fetuses (gestational age≤24 wk, n=5), preterms (gestational age≥25 and ≤36 wk, n=39), and full-term newborns (gestational age≥37 wk, n=18). The average podocyte number of 1,908±645, 1,394±498, and 1,126±256 was, respectively, observed in fetuses, preterms, and full-term newborns. A significant main effect (P=0.0051) of gestational age on podocyte number was observed with a significantly lower number in full-term newborns than in fetuses (P<0.01). Intragroup variability was also observed. We speculate that variations in podocyte number could be correlated with factors such as drugs and maternal diet occurring during intrauterine life. In conclusion, this study shows, for the first time, a decreasing trend in podocyte number during gestation.

Cellular Trafficking of Thymosin Beta-4 in HEPG2 Cells Following Serum Starvation.

Thymosin beta-4 (Tβ4) is an ubiquitous multi-functional regenerative peptide, related to many critical biological processes, with a dynamic and flexible conformation which may influence its functions and its subcellular distribution. For these reasons, the intracellular localization and trafficking of Tβ4 is still not completely defined and is still under investigation in in vivo as well as in vitro studies. In the current study we used HepG2 cells, a human hepatoma cell line; cells growing in normal conditions with fetal bovine serum expressed high levels of Tβ4, restricted to the cytoplasm until 72 h. At 84 h, a diffuse Tβ4 cytoplasmic immunostaining shifted to a focal perinuclear and nuclear reactivity. In the absence of serum, nuclear reactivity was localized in small granules, evenly dispersed throughout the entire nuclear envelop, and was observed as earlier as at 48 h. Cytoplasmic immunostaining for Tβ4 in HepG2 cells under starvation appeared significantly lower at 48 h and decreased progressively at 72 and at 84 h. At these time points, the decrease in cytoplasmic staining was associated with a progressive increase in nuclear reactivity, suggesting a possible translocation of the peptide from the cytoplasm to the nuclear membrane. The normal immunocytochemical pattern was restored when culture cells submitted to starvation for 84 h received a new complete medium for 48 h. Mass spectrometry analysis, performed on the nuclear and cytosolic fractions of HepG2 growing with and without serum, showed that Tβ4 was detectable only in the cytosolic and not in the intranuclear fraction. These data suggest that Tβ4 is able to translocate from different cytoplasmic domains to the nuclear membrane and back, based on different stress conditions within the cell. The punctuate pattern of nuclear Tβ4 immunostaining associated with Tβ4 absence in the nucleoplasm suggest that this peptide might be localized in the nuclear pores, where it could regulate the pore permeability.

Peri and intraglomerular haematoxylinophilic deposits in a newborn: question

A female preterm infant, weighing 2,480 g, was born in a first level Hospital, with caesarean section performed for acute fetal distress. The mother, a 34 years old woman, had a previous healthy child. At birth the general conditions were good. At cardiological examination a patent ductus arteriosus was diagnosed. Laboratory values were within normal range. The newborn was discharged after 11 days, in good general conditions and with spontaneous feeding. During a planned cardiologic follow-up control at day 50, cardiac ultrasounds revealed an increase of volume of left atrium and ventriculus, which appears severely hypokinetic. The baby was urgently transferred in a NICU, where she died after 12 hours for heart insufficiency. At autopsy, deposition of amorphous material, mainly stained by haematoxylin, in periglomerular location was found in the kidney. 1. What is your morphological diagnosis? 2. Which complementary investigations do you suggest? 3. Could be important the examination of large and medium-sized arteries in the heart?

Polyarteritis nodosa in a 21-month-old child: answer

Childhood polyarteritis nodosa (PAN) is a necrotizing vasculitis, affecting small and medium size blood vessels. This condition was first described by Kussmaul and Maier in 1866. Although comparatively rare in childhood, it is the most common form of systemic vasculitis in children. PAN includes two different subtypes, the classical systemic form presenting with a wide range of clinical manifestations including dermatologic, musculoskeletal, nervous, renal, and gastrointestinal systems and the more frequent cutaneous form (CPAN) that involves only the skin. The main clinical features of PAN are malaise, fever, weight loss, skin rash, myalgia, abdominal pain and arthropathy. Systemic involvement is variable, but the skin, the musculoskeletal system, the kidneys and the gastrointestinal tract are most prominently affected, with cardiac, neurological and respiratory manifestations occurring less frequently. However, clinical manifestations can be very confusing, with absence of conclusive diagnostic evidence in the early phase and sometimes in the late phase of the illness. The etiology of PAN remains unclear, but there are data to support roles for hepatitis B and reports of a higher frequency of exposure to parvovirus B19 and cytomegalovirus in PAN patients compared with control populations. However, in childhood, associations between PAN and these infections or other conditions are rare. Evidence has emerged suggesting that bacterial superantigens may play a role in some cases. Here we report the clinico-pathological findings of a 21-month-old child affected with PAN, with particular emphasis on the severity of renal pathological lesions.

Severe renal damage in a 21-month-old child: question

A male child aged 21 months was admitted fifteen days after abdominal pain and vomits. At clinical examination, he had a pale skin, erythematous eruptions on the arms and abdomen, edema of the eyelids and feet, whitish patches on the oral mucosa of the cheek, hypertrophic tonsils and latero cervical lymphadenopathy. Laboratory tests were within the normal range. The patient was not febrile. A few days later, the child’s general condition worsened and fever appeared. Seven days after admission, the child showed a skin rash on the face. He received several drugs, including aspirin and antibiotics. The patient general condition continued to worsen (body weight decreased, blood pressure and fever increased) until death. 1. Which is your macroscopic diagnosis? 2. Which is your histological diagnosis?