Anna Zuk

Transformations Between Epithelium and Mesenchyme: Normal, Pathological, and Experimentally Induced

In this review, we define the two major tissue types, epithelium and mesenchyme, and we describe the transformations (transdifferentiations) of epithelium to mesenchyme (EMT) and mesenchyme to epithelium (MET) that occur during embryonic development. The differentiation of the metanephric blastema provides a striking example of MET. Differentiation of metanephric epithelium is promoted by matrix molecules and receptors (nidogen, laminins, alpha 6 integrins), hepatic growth factor/scatter factor, and products of the genes wnt-1, wnt-4, and Pax-2. Transformation of MDCK epithelium to mesenchyme-like cells is promoted in vitro by antibodies to E-cadherin, products of v-src, v-ras, and v-mos, and by manipulation of the epithelium on collagen gels. Suspension in collagen gel, transforming growth factors, and c-fos have also been shown to promote EMT in epithelia. We present studies from our laboratory showing that alpha 5 beta 1 integrin has a role in the EMT of lens epithelium that is brought about by suspension in collagen gel. Our laboratory has also shown that transfection with the E-cadherin gene induces embryonic corneal fibroblasts to undergo MET and that this MET is enhanced by interaction of the differentiating epithelium with living fibroblasts. This review calls attention to the roles that EMT and MET might have in kidney pathologies and urges further study of the involvement of these phenomena in renal development, renal injury, and renal malignancy.

Polarity, integrin, and extracellular matrix dynamics in the postischemic rat kidney

Acute renal failure (ARF) as a consequence of ischemic injury is a common disease affecting 5% of the hospitalized population. Despite the fact that mortality from ARF is high, there has been little improvement in survival rates over the last 40 years. The pathogenesis of ARF may be related to substantial changes in cell-cell and cell-extracellular matrix interactions mediated by beta1-integrins. On the basis of in vitro and in vivo studies, reorganization of beta1-integrins from basal to apical surfaces of injured tubular epithelia has been suggested to facilitate epithelial detachment, contributing to tubular obstruction and backleak of glomerular filtrate. In this study, we examine integrin and extracellular matrix dynamics during epithelial injury and repair using an in vivo rat model of unilateral ischemia. We find that, soon after reperfusion, beta1-integrins newly appear on lateral borders in epithelial cells of the S3 segment but are not on the apical surface. At later times, as further injury and regeneration coordinately occur, epithelia adherent to the basement membrane localize beta1 predominantly to basal surfaces even while the polarity of other marker proteins is lost. At the same time, amorphous material consisting of depolarized exfoliated cells fills the luminal space. Notably, beta1-integrins are not detected on exfoliated cells. A novel finding is the presence of fibronectin, a glycoprotein of plasma and the renal interstitium, in tubular spaces of the distal nephron and to a lesser extent S3 segments. These results indicate that beta1-integrins dramatically change their distribution during ischemic injury and epithelial repair, possibly contributing to cell exfoliation initially and to epithelial regeneration at later stages. Together with the appearance of large amounts of fibronectin in tubular lumens, these alterations may play a significant role in the pathophysiology of ARF.

Acute Kidney Injury

Acute kidney injury (AKI) is a global public health concern associated with high morbidity, mortality, and healthcare costs. Other than dialysis, no therapeutic interventions reliably improve survival, limit injury, or speed recovery. Despite recognized shortcomings of in vivo animal models, the underlying pathophysiology of AKI and its consequence, chronic kidney disease (CKD), is rich with biological targets. We review recent findings relating to the renal vasculature and cellular stress responses, primarily the intersection of the unfolded protein response, mitochondrial dysfunction, autophagy, and the innate immune response. Maladaptive repair mechanisms that persist following the acute phase promote inflammation and fibrosis in the chronic phase. Here macrophages, growth-arrested tubular epithelial cells, the endothelium, and surrounding pericytes are key players in the progression to chronic disease. Better understanding of these complex interacting pathophysiological mechanisms, their relative importance in humans, and the utility of biomarkers will lead to therapeutic strategies to prevent and treat AKI or impede progression to CKD or end-stage renal disease (ESRD).

Induction of the endogenous whey acidic protein (Wap) gene and a Wap-myc hybrid gene in primary murine mammary organoids.

In rodents, the whey acidic protein (Wap) is the major whey protein expressed in mammary glands in response to lactogenic hormones. The regulation of the Wap gene differs from that of other milk protein genes, with one consequence being that little or no Wap expression is detectable in cell culture. Here we describe the efficient in vitro induction of the Wap gene in mammary organoids isolated from midpregnant mice. Mammary organoids were isolated as intact epithelial subcomponents which retained the glandular microarchitecture. If organoids were cultured in contact with a monolayer of 3T3-L1 adipocytes, significant levels of Wap mRNA were induced upon hormonal stimulation, with the highest level of Wap mRNA being induced by a combination of hydrocortisone, prolactin, and insulin. Dissociation of the three-dimensional organization abrogated Wap inducibility. Organoids cultured on plastic or hydrated type I collagen did not transcribe Wap mRNA even after hormonal stimulation. Addition of hormones was required to maintain low levels of Wap mRNA in organoids cultured on reconstituted basement membrane, however, Wap mRNA was not induced. Organoid-adipocyte interactions as well as cell-cell interactions inherent in the structure of organoids promote hormone-dependent Wap mRNA expression. In order to study the Wap promoter region in vitro, we cocultured organoids from transgenic mice harboring a chimeric Wap-myc gene with 3T3-L1 adipocytes. Lactogenic hormones induced the Wap-myc transgene in vitro. The kinetics of induction were similar for both the transgene and the endogenous Wap gene indicating that the 2.5-kb regulatory Wap region present in the hybrid gene contains the sequence elements required for hormone-induced gene expression in vitro.

Integrins in epithelial cell polarity: Using antibodies to analyze adhesive function and morphogenesis

Epithelial cells polarize in response to cell-substratum and cell-cell adhesive interactions. Contacts between cells and proteins of the extracellular matrix are mediated by integrin receptors. Of the 24 recognized integrin heterodimers, epithelial cells typically express four or more distinct integrins, with the exact complement dependent on the tissue of origin. Investigation of the roles of integrins in epithelial cell polarization has depended on the use of function-blocking antibodies both to determine ligand specificity of individual integrins and to disrupt and redirect normal morphogenesis. In this article we describe techniques for employing function-blocking anti-integrin antibodies in adhesion assays of the polarized Madin-Darby canine kidney (MDCK) cell line and to demonstrate the involvement of beta1 integrins in collagen-induced tubulocyst formation. These techniques can be easily expanded to other antibodies and epithelial cell lines to characterize specific functions of individual integrins in epithelial morphogenesis.

Acute kidney injury: Can remote ischaemic preconditioning prevent AKI?

A randomized multicentre controlled study of 240 cardiac surgery patients at high risk of acute kidney injury (AKI) has demonstrated that remote ischaemic preconditioning can reduce the rate of AKI and requirement for renal replacement therapy. These findings suggest this procedure could be a promising therapeutic option for this high-risk patient group.

Can remote ischaemic preconditioning prevent AKI

A randomized multicentre controlled study of 240 cardiac surgery patients at high risk of acute kidney injury (AKI) has demonstrated that remote ischaemic preconditioning can reduce the rate of AKI and requirement for renal replacement therapy. These findings suggest this procedure could be a promising therapeutic option for this high-risk patient group.

Can remote ischaemic preconditioning prevent AKI?: Acute kidney injury

A randomized multicentre controlled study of 240 cardiac surgery patients at high risk of acute kidney injury (AKI) has demonstrated that remote ischaemic preconditioning can reduce the rate of AKI and requirement for renal replacement therapy. These findings suggest this procedure could be a promising therapeutic option for this high-risk patient group.