The loss of expression of transforming growth factor-beta receptors correlates with the histopathologic tumor grade in bladder transitional cell carcinoma patients.

Transforming growth factor-beta (TGF-beta), a pleiotropic growth factor, is a potent inhibitor of cellular proliferation in cells of epithelial origin. Recently, it has been suggested that a loss of sensitivity to TGF-beta through a loss of expression of TGF-beta receptors T beta R-I and T beta R-II--is associated with tumor initiation and progression. Therefore, to investigate the relationship between TGF-beta receptors expression and carcinogenesis of bladder TCC, this study examined the expression of T beta R-I and T beta R-II in 46 bladder TCC patients using immunohistochemistry. Since histopathological grade is a widely accepted marker of prognosis, the results were compared in relation to the three grades of bladder TCC. The results demonstrated that the loss of TGF-beta receptors expression is associated with increasing histopathological grades of bladder TCC. Specifically, both T beta R-I and T beta R-II were readily detected in all 10 normal bladder mucosa specimens. Likewise, all 6 specimens of grade I TCC samples expressed high levels of both TGF-beta receptors. However, among grade II TCC samples, T beta R-I and T beta R-II were detected in 78% and 89%, respectively: among grade III TCC samples, T beta R-I and T beta R-II were detected in 45% and 41%, respectively. These results suggested that loss of sensitivity to TGF-beta may play a role in the progression of TCC from low to high grade disease.     

Expression of protein kinase C genes in normal (+/+) and W mutant alleles (Wsh/Wsh, W/Wv) mice testes

In the present study, we investigated the expression of mRNA of protein kinase C (PKC) isoenzymes (alpha, beta, gamma, delta, epsilon, zeta, eta, and theta) in normal (+/+) and W mutant alleles mice testes. In +/+ mice testes, abundant expression of PKCdelta and PKCtheta was observed, while other PKCs (alpha, beta, gamma, epsilon, zeta, and eta) generally were not detected by Northern blotting. The PKCdelta and PKCtheta isoenzymes demonstrated a distinctive cellular distribution when evaluated by in situ hybridization. We have previously shown that PKCdelta gene was selectively expressed in spermatid of +/+ testes. Here we show that PKCdelta gene is also present in spermatid of Wsh/Wsh mice testes and PKCtheta gene was present in interstitial cells of +/+, Wsh/Wsh, and W/Wv mice testes. These studies provide the evidence of selective cell distributions of the PKC isoenzymes and suggest that PKC has the functional significance in testes.     

Asbestos-induced apoptosis is protein kinase C delta-dependent

Alveolar epithelial and mesothelial cells undergo apoptosis in response to asbestos, a phenomenon that may be important in injury and/or initiation of compensatory proliferation. Here, we report a functional role of protein kinase (PKC)delta in apoptosis by crocidolite asbestos. We first show that asbestos increases the kinase activity of PKC delta in alveolar type II epithelial cells (C10 line) and causes its translocation to mitochondria, events associated with caspase-9 cleavage and apoptosis as detected by the Apostain technique. Pretreatment of C10 cells with rottlerin (Rot), a PKC delta-selective inhibitor, before addition of asbestos prevented cleavage of caspase-9 and blocked the appearance of apoptotic cells. Asbestos-induced apoptosis also was inhibited in cells stably expressing a dominant-negative kinase-deficient mutant of PKC delta (dnPKC delta), but not dnPKC alpha. Activities of PKC alpha and PKC zeta increased after exposure to asbestos, but neither isoform migrated to mitochondria. A general inhibitor of PKCs, bisindolylmaleimide I, had no effect on asbestos-induced apoptosis. Hydrogen peroxide (H2O2) induced activation of PKCs delta, alpha, zeta, and theta, translocation of PKC delta to mitochondria, and caspase-9 cleavage. However, H2O2-induced apoptosis was not inhibited by cell lines stably expressing either dnPKC delta or dnPKC alpha, suggesting that activation of PKC delta has a distinct role in the development of asbestos-induced apoptosis.     

Expression of transforming growth factor beta1 and its receptors in normal human urothelium and human transitional cell carcinomas

Previous studies indicated that transforming growth factor beta1 (TGFbeta1) is expressed by normal urothelial cells and exerts regulatory autocrine functions in urothelial maintenance and wound healing. However, little is known about the expression patterns of TGFbeta1 and its receptors in bladder tumors. Therefore, we studied the protein and mRNA localization of TGFbeta1 and TGFbeta receptor types I and II (TGFbetaRI and TGFbetaRII) in normal human urothelium and transitional cell carcinomas (TCCs) of different grades and stages. Expression of TGFbeta1 and its receptors was examined by immunocytochemistry and mRNA in situ hybridization in normal urothelium and TCCs using a semiquantitative method. By immunocytochemistry, the expression of TGFbeta1 and TGFbetaRII was higher in superficial and basal cell layers of normal urothelium than in the intermediate layer. A similar localization was seen in superficial TCCs. TGFbetaRI was mainly present in basal and intermediate cell layers of normal urothelium and superficial TCCs. In contrast, in muscle invasive TCCs, all tumor cells stained intensely for all three proteins. No correlation was found between immunostaining and TCC grade. In situ hybridization pointed out that all cell layers in normal urothelium exhibit similar TGFbeta1 mRNA levels. Elevated TGFbeta1 mRNA levels were noted in TCCs irrespective of grade or stage. In conclusion, these data indicate that in normal urothelium TGFbeta1, TGFbetaRI, and TGFbetaRII expression depend on maturation and differentiation. This pattern is particularly lost in muscle invasive TCCs, in which the expression of the three proteins is enhanced. These data suggest autocrine TGFbeta1 mechanisms in human TCC cells that may be more pronounced in muscle invasive TCC cells.     

Expression of the VLA beta 1 integrin family in bladder cancer.

Integrins are a family of transmembrane heterodimers, many of which function as receptors for extracellular matrix molecules and play a role in adherence to and motility on matrix components. Because of these functions, integrins are suspected of participating in metastatic processes. We investigated the expression of beta 1 integrins in human bladder cancer cell lines and tissues. Expression of beta 1 integrins on cultured bladder cancer cell lines was evaluated by flow cytometry, of 8 cell lines tested, alpha 1 was found in 4, alpha 2 and alpha 3 in all 8, alpha 4 in 1, and alpha 5 in 3. These results were in sharp contrast to the expression detected by immunostaining tissues containing normal urothelium and low stage (noninvasive) and high stage (invasive) bladder cancers. All normal urothelial tissues tested expressed alpha 2 and alpha 3 and none expressed alpha 1, alpha 4, or alpha 5. Similarly, a majority (77%) of low stage (noninvasive) bladder cancers stained positively for alpha 3, whereas only 6 of 13 expressed alpha 2 and none expressed alpha 1, alpha 4, or alpha 5. Among invasive bladder cancers, alpha 1 was detected in 7%, alpha 2 in 24%, alpha 3 in 68%, alpha 5 in 10%, and alpha 4 was not found in any samples. These results indicate that integrin expression in cultured human bladder cancer cell lines does not represent expression observed in tissue samples and may reflect adaption to or selection during tissue culture conditions. A progressive loss of alpha 2 expression is seen from normal urothelial cells through invasive bladder cancers. This loss may contribute to an invasive phenotype by a loss of the cell-cell adherence function mediated by the alpha 2 beta 1 and alpha 3 beta 1 integrins.     

Caveolin-1 Is Down-Regulated in Human Ovarian Carcinoma and Acts as a Candidate Tumor Suppressor Gene

To identify novel markers differentially expressed in ovarian cancer versus normal ovary, we hybridized microarrays with cDNAs derived from normal human ovaries and advanced stage ovarian carcinomas. This analysis revealed down-regulation of the caveolin-1 gene (CAV1) in ovarian carcinoma samples. Suppression of CAV1 in ovarian carcinomas was confirmed using a tumor tissue array consisting of 68 cDNA pools from different matched human tumor and normal tissues. Immunohistochemistry demonstrated expression of caveolin-1 in normal and benign ovarian epithelial cells, but loss of expression in serous ovarian carcinomas. In low-grade carcinomas, redistribution of caveolin-1 from a membrane-associated pattern observed in normal epithelium to a cytoplasmic localization pattern was observed. No expression of caveolin-1 was detectable in four of six ovarian carcinoma cell lines investigated. In SKOV-3 and ES-2 carcinoma cells, which express high levels of the caveolin-1 protein, phosphorylation of the 22-kd caveolin-1 isoform was detected. Inhibition of both DNA methylation and histone deacetylation using 5-aza-2'deoxycytidine and Trichostatin A, respectively, relieves down-regulation of caveolin-1 in OAW42 and OVCAR-3 cells which is in part mediated by direct regulation at the mRNA level. Expression of CAV1 in the ovarian carcinoma cell line OVCAR-3, resulted in suppression of tumor cell survival in vitro, suggesting that the CAV1 gene is likely to act as a tumor suppressor gene in human ovarian epithelium.     

Regulation of cell surface expression of Fas (CD95) ligand and susceptibility to Fas (CD95)-mediated apoptosis in activation-induced T cell death involves calcineurin and protein kinase C, respectively

We show that an influenza hemagglutinin-specific CD4+ murine T cell hybridoma (IP-12-7) enters the apoptotic suicide program via the Fas ligand (FasL)/Fas-mediated pathway upon T cell receptor (TCR) stimulation. These cells express Fas and FasL mRNA, cell surface Fas and intracellular FasL, but do not enter apoptosis upon Fas ligation prior to TCR stimulation. TCR stimulation additionally results in protein synthesis-dependent cell surface expression of the preformed FasL. Addition of phorbol dibutyrate (PBu2) alone was sufficient to induce susceptibility to Fas ligation induced apoptosis, while addition of both PBu2 and calcium ionophore A23187 were required to induce FasL cell surface expression. Addition of cyclosporin A completely inhibited TCR-mediated death and FasL cell surface up-regulation, but had no effect on apoptosis induced directly by Fas ligation following TCR stimulation. Inhibitors of protein kinase C (PKC) (G? 6976 and GF 109203X) completely inhibited TCR-induced susceptibility to Fas ligation, but only partially inhibited TCR-induced cell surface expression of FasL. PKC isoenzymes alpha, beta, delta and zeta were expressed by this cell line and only the alpha and betaI isoforms translocated to the membrane fraction upon TCR stimulation. Our data suggest that in activation-induced T cell apoptosis PKC is involved in pathways that mediate the acquisition of Fas susceptibility, while calcineurin is required for cell surface expression of the preformed FasL.     

Evaluation of chromosome 8 and 11 aneuploidies in washings and biopsy materials of bladder transitional cell carcinoma

We compared chromosome 8 and 11 aneuploidies on bladder biopsy tumor tissues and bladder washing samples of transitional cell carcinoma (TCC) and their relationship to tumor malignancy. Interphase fluorescence in situ hybridization (FISH) was applied to nuclei of washing material and biopsy samples of 17 patients with TCC. Incidence of cells having aneuploidy was clearly nonrandom from patient to patient. There was no significant difference in the incidence of aneuploid frequency for chromosomes 8 and 11 between biopsies of bladder tumors and bladder washing samples (P > 0.05). For chromosome 8, incidence of disomic cells (having two signals) in grade III tumors was significantly lower than in grade II tumors of both washing samples (P = 0.004) and biopsy materials (P = 0.005), indicating a high frequency of aneuploidy. The incidence of nuclei with four or more than four signals of chromosome 8 was significantly higher in grade III tumors than in grade II tumors in washing samples (P = 0.031 and 0.003, respectively). Similarly, in biopsy material, the incidence of nuclei with more than four signals of chromosome 8 was significantly higher in grade III tumors than in grade II tumors (P = 0.004). For chromosome 11, in both washing samples and biopsy materials, the incidence of disomic cells (having two signals) in grade III tumors was significantly lower than that detected in grade II tumors (P = 0.031 and 0.014, respectively), indicating a high frequency of aneuploidy. In biopsy materials, the incidence of nuclei with three or four signals was significantly higher than that in grade II tumors (P = 0.014 and 0.012, respectively). These findings suggest that FISH analysis of bladder washing samples can be effectively detected as genetic changes of bladder tumors. It might predict genetic progression of these tumors, which might be related to tumor stage, because higher stages of tumors showed a higher incidence of aneuploidies of chromosomes 8 and 11.     

Analysis of protein kinase C delta (PKC delta) expression in endometrial tumors

Endometrial cancer is the most common gynecologic malignancy in the United States. However, its underlying molecular mechanisms are poorly understood; and few prognostic indicators have been identified. The protein kinase C (PKC) family has been shown to regulate pathways critical to malignant transformation; and in endometrial tumors, changes in PKC expression and activity have been linked to a more aggressive phenotype and poor prognosis. We have recently shown that PKC delta is a critical regulator of apoptosis and cell survival in endometrial cancer cells; however, PKC delta levels in endometrial tumors had not been determined. We used immunohistochemistry to examine PKC delta protein levels in normal endometrium and endometrioid carcinomas of increasing grade. Normal endometrium exhibited abundant nuclear and cytoplasmic staining of PKC delta confined to glandular epithelium. In endometrial tumors, decreased PKC delta expression, both in intensity and fraction of epithelial cells stained, was observed with increasing tumor grade, with PKC delta being preferentially lost from the nucleus. Consistent with these observations, endometrial cancer cell lines derived from poorly differentiated tumors exhibited reduced PKC delta levels relative to well-differentiated lines. Treatment of endometrial cancer cells with etoposide resulted in a translocation of PKC delta from cytoplasm to nucleus concomitant with induction of apoptosis. Decreased PKC delta expression, particularly in the nucleus, may compromise the ability of cells to undergo apoptosis, perhaps conferring resistance to chemotherapy. Our results indicate that loss of PKC delta is an indicator of endometrial malignancy and increasing grade of cancer. Thus, PKC delta may function as a tumor suppressor in endometrial cancer.     

Characterization of expression of protein kinase C isozymes in human B-cell lymphoma: Relationship between its expression and prognosis

Protein kinase C (PKC) enzymes play a major role in signal transduction and contribute to the regulation of cellular differentiation and proliferation. However, little is known about subtype-specific intracellular expression of PKC in human malignant lymphoma. To characterize the relationship between expression of PKC and B-cell lymphomas based on the different subspecies, we investigated the expression of four subspecies (alpha, beta II, gamma and delta) in five cases of reactive lymphoid tissues, 77 cases of human B-cell lymphoma and 17 human lymphoma cell lines. In the reactive lymphoid tissues, PKC beta II-positive cells were found in the mantle zones and marginal zones, and centroblasts and centrocytes in the germinal centers showed cytoplasmic staining with strong intensity against PKC delta. The present study is the first report to examine the expression of PKC delta in reactive lymphoid tissues. In interfollicular areas, a small number of T-cells were positive for PKC alpha. Protein kinase C gamma-positive cells were not found in these lymphoid tissues. Eight cases of Burkitt lymphoma (BL) (8/10; 80%) showed the overexpression of PKC alpha (P < 0.01), but other B-cell lymphoma cases except three cases of diffuse large B-cell lymphoma did not express PKC alpha. In addition, six and eight out of nine BL cell lines expressed the protein and mRNA of PKC alpha, respectively. These results indicate that PKC alpha was predominantly expressed on BL in comparison with other types of lymphoma. The expression of PKC gamma was observed in only five cases of BL. The overall survival of PKC gamma-positive BL was significantly better than that of PKC gamma-negative BL (P < 0.05). The expression of PKC gamma seems to be associated with a better prognosis in the limited number of BL cases in the present study.     

Immunocytochemical expression of protein kinase C isoenzymes alpha, delta, epsilon and zeta in differentiating chick chondrocytes in vitro

In our previous work we have investigated the expression of the serine-threonine kinase protein kinase C (PKC) in the vertebral column of mouse foetuses. In the present work we would verify the expression of four PKC-isoenzymes (alpha, delta, epsilon, zeta) in two distinct phases of the chondrogenesis and the endochondral osteogenesis in vitro. We performed primary cultures of chondrocytes collected from tibiae of 6-day old chick embryos. This cells were cultured for 20 days and than collected on coverslips (stage 1 culture). Other cells of the stage 1 were undergone further differentiation towards the phenotype of osteoblast-like cells (stage 2 culture), in accord to the protocol of Descalzi Cancedda et al. (1992). In stage 1 culture, PKC-epsilon was the most expressed isoform, whereas PKC-alpha exhibited the least intense positivity. In stage 2 culture, PKC-alpha was the most expressed isoform, whereas a marked decrease of PKC-epsilon expression was detected compared to stage 1. No relevant differences were evidenced as regards,the expression of PKC-zeta between the two considered cell culture stages. On these bases, it could be reasonable that these PKC-isoenzymes may be involved at different levels in chondrocytes differentiation as well as in the endochondral ossification process.     

VEGF/VEGFR在膀胱癌中表达的研究

目的：探讨血管内皮细胞生长因子（VEGF）和血管内皮细胞生长因子受体（VEGFR）KDR在膀胱移行细胞癌（TCC）患者的组织标本及正常膀胱黏膜组织中的定位及表达情况.方法：分别采用免疫组化SABC法和TUNEL法检测60例膀胱癌组织,并以40例正常膀胱黏膜组织作为对照,比较两种不同组织中VEGF及KDR的阳性表达率和表达强度的差异.结果：在60例膀胱TCC中,VEGF和KDR分别有53例和51例呈阳性表达,平均表达率分别为88%和85%,随肿瘤病理分期和细胞分级的增高其表达水平上调.但在40例正常对照组中无一例表达.两者分别比较,差异均极显著（P＜0.01）.结论：VEGF可能通过膀胱移行细胞上的相应受体而发挥一定的生物学作用.膀胱TCC患者的肿瘤组织中KDR的表达可能直接诱发了肿瘤血管的形成.     

Differentiation-dependent switch in protein kinase C isoenzyme activation by FcgammaRI, the human high-affinity receptor for immunoglobulin G

Aggregation of receptors for the constant region (Fc) of immunoglobulin G on myeloid cells results in endocytosis or phagocytosis and cellular activation. Previous work has shown, using the cell line U937, that the high-affinity immunoglobulin G receptor, FcgammaRI, activates alternate intracellular signalling pathways depending on the cell differentiation state, which results in a marked change in the nature of calcium transients within the cell. Here, we show that protein kinase C (PKC) is activated in both interferon-gamma (IFN-gamma) -primed and dibutyryl cyclic AMP (dbcAMP) -differentiated cells but that the nature of the particular isoenzymes recruited differs. Thus, in IFN-gamma-primed U937 cells, FcgammaRI aggregation results in an increase of PKC activity which is essentially calcium independent resulting from the translocation to the membrane of the novel PKCs, delta and epsilon, together with the atypical PKC zeta. However, in cells differentiated to a more macrophage phenotype, all PKC enzyme activity after receptor aggregation is calcium dependent. Consistent with this finding, the isoenzymes translocated to the nuclear-free membrane fraction are the conventional PKCs alpha, beta and gamma; results consistent with our previous finding that FcgammaRI couples to phospholipase C in such dbcAMP-differentiated cells. Thus, the nature of PKC isoenzyme activated following FcgammaRI aggregation is defined by differentiation. The calcium dependence of the PKC isoenzyme is consistent with the duration of calcium transients previously reported in the two differentiation states.     

Genetic and phenotypic changes associated with the acquisition of tumorigenicity in human bladder cancer

There has been a general lack of human paired cell lines that both reproduce the in vivo spectrum of tumor progression of bladder cancer and have some of the genetic changes associated with progression in human tumor tissue. T24, a cell line established from an invasive human transitional cell carcinoma (TCC) of the bladder, has been used extensively in bladder cancer research. However, a significant limitation of this cell line is its lack of tumorigenicity when injected into immunocompromised mice. This characteristic was used to our advantage as we sought to characterize T24T, a highly tumorigenic variant that could then be used to elucidate the genes responsible for human bladder tumor progression. In culture, T24T has a faster doubling time, reaches a higher cell density in monolayer culture, and is more motile than T24 at higher cell densities. T24T is able to form colonies in soft agar, whereas T24 is not, and expresses HRAS, a gene associated with increased aggressiveness in human TCC, at higher levels than T24. Most importantly, T24T forms solid tumors when injected subcutaneously in SCID mice both with and without Matrigel (Sigma, St. Louis, MO), whereas T24 does not. Cytogenetically, the 2 cell lines contain at least 5 shared structural anomalies, as determined by detailed karyotyping. Interestingly, T24T has acquired 4 new structural changes, 3 of which [add(10)(p12), i(10)(q10), -15] have been observed in loss of heterozygosity (LOH) studies of tumor progression in human TCC. It appears that the T24/T24T model may be an excellent tool for the study of human TCC progression because of its relationship with known karyotypic changes associated with human bladder cancer progression. We are currently taking advantage of these paired cell lines to identify genes involved in human TCC progression. Genes Chromosomes Cancer 27:252-263, 2000.     

Human urinary bladder transitional cell carcinomas acquire the functional Fas ligand during tumor progression

The interaction between FasL on tumor cells and Fas on lymphocytes may represent a tumor immune escape mechanism. We explored FasL expression and function in human urinary bladder transitional cell carcinomas (TCCs). FasL expression was observed in situ in 45% of TCCs (n = 45) and was absent in normal urothelium (n = 20). A correlation existed between FasL expression and high tumor grade (0% in G1, 14% in G2, and 75% in G3; P < 0.0001) and stage (13% in superficial Ta-T1 versus 81% in invasive T2-T4; P < 0.0001). FasL function was shown by the ability of two FasL-positive primary culture TCC cell lines (established from two FasL-positive invasive TCCs) to induce Fas-mediated killing not only of conventional Fas-sensitive targets (such as Jurkat cells or phytohemagglutinin-lymphoblasts), but also of autologous T lymphocytes generated in a mixed lymphocyte tumor-cell culture. In addition, an association between FasL expression by TCC cells and activated caspase-8, -9, and -3 expression by interferon-gamma-producing CD8-positive tumor-infiltrating lymphocytes was observed in situ. Our results show a functional expression of TCC-expressed FasL that correlates with tumor progression. These results suggest that TCC-expressed FasL may induce apoptosis of anti-tumor T lymphocytes in vivo, providing new insights on the mechanisms involved in bladder TCC progression.     

Expression of bcl-2 in bladder neoplasms is a cell lineage associated and p53-independent event.

AIMS: To investigate bcl-2 and p53 protein expression in hyperplastic, metaplastic and neoplastic epithelia of the urinary bladder in relation to cell lineages (transitional versus glandular epithelia). METHODS: Formalin fixed, paraffin wax embedded archival tissue blocks of 29 transitional cell carcinomas (TCC), 11 adenocarcinomas, five specimens of cystitis glandularis, four papillomas, and seven samples of morphologically normal bladder mucosa were examined immunohistochemically with antibodies specific to bcl-2 and p53. Consecutive sections were used to assess co-expression of the two proteins. RESULTS: bcl-2 protein was expressed heterogeneously in basal cells of the normal transitional epithelium, whereas p53 was rarely detectable in either normal or hyperplastic epithelium. Of the 29 TCCs, 20 (69%) expressed immunodetectable p53 which was positively associated with grade. In contrast, bcl-2 was detected in four (14%) TCCs and its expression was not associated with grade. bcl-2 was expressed constitutively in all five specimens of cystitis glandularis and in all adenocarcinomas; p53 was co-expressed in most of the latter. There was no association between bcl-2 and p53 protein expression in the TCCs. Expression of bcl-2 protein correlated negatively with grade of adenocarcinoma. CONCLUSION: In bladder adenocarcinomas, bcl-2 expression correlated negatively with tumour grade whereas p53 was associated positively with tumour grade. The association of bcl-2 with cystitis glandularis and adenocarcinoma but not TCC suggests that it may be involved in triggering a lineage switch converting transitional epithelium to a glandular phenotype.     

Use of monoclonal antibody E48 in diagnosing transitional cell carcinoma of urinary bladder.

AIMS: To investigate the localisation of the E48 epitope and to determine the use of monoclonal antibody E48 for the identification of transitional cell carcinomas (TCC); and to determine if antigenic expression was affected by different standard fixation methods. METHODS: Biopsy specimens were labelled with E48 for immunoelectron microscopy. One hundred and nineteen tissue samples from 47 bladder carcinomas were tested for reactivity with E48, using fresh frozen, sublimate formalin, and formalin fixed tissue. Thirteen undifferentiated bladder tumours and 10 undifferentiated prostatic carcinomas were incubated with E48 and prostate specific antigen. RESULTS: Reactivity to E48 was found in all grade 1 and 2 carcinomas and most (83%) grade 3 tumours. At the ultrastructural level, expression was mainly associated with desmosomes and the cytoplasmic membrane. The reactivity of E48 was generally strong in fresh frozen tissue samples and remained preserved in fixed tissue samples. Ten of the 13 bladder carcinomas expressed E48; all prostatic tumours were totally negative. CONCLUSIONS: E48 is a sensitive marker for transitional cell carcinoma and suitable for differentiation between urothelial and prostatic undifferentiated carcinoma. It can be used in routinely processed, formalin fixed, biopsy specimens.     

An atypical protein kinase C, PKC zeta, regulates human eosinophil effector functions

Protein kinase (PK) C comprises a family of isoenzymes that play key roles in downstream signalling and cell functions. We studied PKC zeta participation in the effector functions of human eosinophils stimulated with platelet-activating factor (PAF) or complement (C) 5a. After pretreating eosinophils with a myristoylated specific PKC zeta inhibitor; bisindlolylmaleimide I (BisI), an inhibitor of conventional and novel PKCs; or rottlerin, a PKC delta inhibitor, we examined PAF- and C5a-evoked functions. Induced PKC translocation was characterized by confocal laser scanning microscopy. The PKC zeta inhibitor blocked PAF- or C5a-induced eosinophil superoxide anion generation as effectively as BisI or rottlerin. The PKC zeta inhibitor also attenuated PAF- or C5a-induced eosinophil degranulation and adhesion. In contrast, the PKC zeta inhibitor did not affect PAF- or C5a-induced CD11b expression. Finally, both eosinophil shape changes and the translocation of PKC zeta and p47phox, a component of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, to the plasma membrane induced by PAF or C5a were completely inhibited by the PKC inhibitor. Thus, the atypical PKC zeta regulates human eosinophil adhesion and effector functions.