Congenital obstructive nephropathy accounts for the greatest fraction of chronic kidney disease in children. Genetic and nongenetic factors responsible for the lesions are largely unidentified, and attention has been focused on minimizing obstructive renal injury and optimizing long-term outcomes. The cellular and molecular events responsible for obstructive injury to the developing kidney have been elucidated from animal models. These have revealed nephron loss through cellular phenotypic transition and cell death, leading to the formation of atubular glomeruli and tubular atrophy. Altered renal expression of growth factors and cytokines, including angiotensin, transforming growth factor-beta, and adhesion molecules, modulate cell death by apoptosis or phenotypic transition of glomerular, tubular, and vascular cells. Mediators of cellular injury include hypoxia, ischemia, and reactive oxygen species, while fibroblasts undergo myofibroblast transformation with increased deposition of extracellular matrix. Progression of the lesions involves interstitial inflammation and interstitial fibrosis, both of which impair growth of the obstructed kidney and result in compensatory growth of the contralateral kidney. The long-term outcome depends on timing and severity of the obstruction and its relief, minimizing ongoing injury, and enhancing remodeling. Advances will depend on new biomarkers to evaluate the severity of obstruction, to determine therapy, and to follow the evolution of lesions.

Following UUO, the co-localization of hypoxia with cellular proliferation, necrosis, and TBM thickening of the PT is consistent with ischemic injury resulting from vasoconstriction. In contrast, a selective dilation of the distal portion of the nephron (DT and CD), which results from the greater tubular compliance there, leads to stretch-induced epithelial cell apoptosis, along with a progressive peritubular fibrosis. Nephron loss in the obstructed developing kidney likely results from complex, segment-specific cellular responses.

We have used an interaction cloning strategy to isolate cDNAs for sequences that interact with protein kinase C (Chapline, C., Ramsay, K., Klauck, T., and Jaken, S. (1993) J. Biol. Chem. 268, 6858 -6861). In this paper, we report a novel sequence, clone 72, isolated according to this method. Clone 72 has a 4.8-kilobase pair open reading frame; antibodies to clone 72 recognize a >200-kDa protein in cell and tissue extracts. Clone 72 message and protein are detected in a variety of tissues. Immunoprecipitation studies demonstrate that clone 72 is the major >200-kDa binding protein described previously in REF52 fibroblasts (Hyatt, S. L., Liao, L., Aderem, A., Nairn, A., and Jaken, S. Protein kinase C (PKC)1 is a family of phospholipid-dependent kinases involved in basic cellular functions, including regulation of growth, differentiation, and gene expression (1, 2). The role of individual PKCs in these processes is not yet known; however, since most cells express more than one type of PKC, it seems likely that individual PKCs have unique rather than overlapping functions. All of the PKCs require phosphatidylserine for maximal activity; however, PKCs can be grouped according to differences in their dependence on other activators. In addition to phosphatidylserine, conventional PKCs require calcium and diacylglycerol, novel PKCs require only diacylglycerol, and atypical PKCs require nothing more. Several other lipid modifiers of PKC activity have also been identified, and there is some evidence that they may selectively influence individual isozyme activities (1). Selective isozyme activation in response to physiological agonists has been noted and may be a result of the recognized differences in cofactor requirements among the PKCs (3-6).In addition to isozyme selective activation, isozyme-specific functions may depend on selective substrate recognition. However, only minor differences in substrate specificity among the isozymes have been observed in in vitro assays (7,8). Recently, immunofluorescence studies have demonstrated unique subcellular localizations for individual PKCs (9 -11) (data not shown). Thus, targeting of individual PKCs to specific subcellular addresses may be a means of restricting accessibility to substrates and, thereby, provide the mechanism for isozyme-selective phosphorylation events in vivo. To isolate proteins that interact with PKCs with high affinity, we developed an assay for identifying PKC-binding proteins (12-14). Subsequently, this assay was adapted to screen expression libraries and isolate cDNA clones for PKC-binding proteins (15). Binding proteins isolated according to this strategy are also substrates (16). In this manuscript, we report the full-length sequence of one of these binding proteins, clone 72. The results indicate that clone 72 is widely expressed and is a major PKC-binding protein in REF52 fibroblasts. MATERIALS AND METHODSLibrary Screening-A gt11 REF52 cDNA library was prepared and screened for PKC-binding proteins as described in Ref. 15. Positive colonies were pla...

Post-transplantation lymphoproliferative disorder (PTLD) is a widely recognized and often catastrophic complication of organ transplantation. The incidence of PTLD after lung transplantation ranges from 6.2 to 9.4% and is two-fold higher than that seen after organ transplantation of other organs. Primary Epstein-Barr virus (EBV) infection is a major risk factor for PTLD, but the incidence of PTLD in EBV seronegative (EBV-) patients seems to vary with type of organ transplant. The goal of this study was to quantify the risk of PLTD based on pre-lung transplantation EBV serostatus in lung transplant patients. Pre- and post-lung transplant serostatus was defined in 80 patients, and our six cases of PTLD occurred in this group. Six of 94 lung transplant patients (6.4%) who survived > 1 mo developed PTLD. All cases of PTLD involved thoracic structures at presentation and occurred in the first post-operative year. Patients who were EBV- before lung transplant were much more likely to develop PTLD than those who were seropositive (EBV+) (five of 15 [33%] versus one of 60 [< 2%], p < 0.001). Consistent with the prevailing adult (donor) EBV+ rate (85%), two of our EBV-patients remained EBV-after lung transplant. Therefore, the rate of PTLD was 42% in those with primary EBV infection. As compared with EBV-patients that remained tumor-free, those who developed PLTD had similar levels of immunosuppressants and doses of anti-viral therapy. We conclude that PLTD occurs predominantly in EBV-naïve patients (risk approximately 1/3). EBV-patients should be monitored more closely after lung transplantation and, possibly, managed with lower immunosuppression. Our data also suggest that anti-viral therapy alone does not decrease the incidence of PTLD in high risk patients, PTLD can be successfully treated in most cases, and EBV-naïve patients should not be excluded from lung transplant because their risk of death from PTLD is < 15%.

Relatively little is known about the molecular mechanisms of tumor promotion/progression in mammary carcinogenesis. Increased protein kinase C (PKC) activity is known to promote tumor formation in several tissues; however, its role in mammary carcinogenesis is not yet known. To determine if individual PKCs may selectively regulate properties of mammary tumor cells, we compared PKC isozyme levels in mammary tumor cell lines with low, moderate and high metastatic potential. All three cell lines expressed a, d, e and z PKCs; however, PKCd levels were relatively increased in the highly metastatic cells. To determine if increased PKCd could contribute to promotion/progression, we overexpressed PKCd in the low and moderately metastatic cell lines. PKCd overexpression had no signi®cant e ect on growth of adherent cells, but signi®cantly increased anchorage-independent growth. Conversely, expressing the regulatory domain of PKCd (RDd), a putative PKCd inhibitory fragment, inhibited anchorage-independent growth. The e cacy of RDd as a PKCd inhibitor was demonstrated by showing that RDd selectively interfered with PKCd subcellular location and signi®cantly interfered with phosphorylation of the PKC cytoskeletal substrate, adducin. PKC-dependent phosphorylation of cytoskeletal substrate proteins, such as adducin, provides a mechanistic link between increased PKCd activity and phenotypic changes in cytoskeletaldependent processes such as migration and attachment, two processes that are relevant to metastatic potential. The reciprocal growth e ects of expressing PKCd and RDd as gain and loss of function constructs, respectively, provide strong evidence that PKCd regulates processes important for anchorage-independent growth in these mammary tumor cells.