Biomedical subjects

Istvan Arany

Publications and source records attributed to Istvan Arany.

11 recordsLinked to original sources

PubMed2005

CREB mediates ERK-induced survival of mouse renal tubular cells after oxidant stress.

BACKGROUND: We showed that extracellular signal-regulated protein kinase (ERK) is prosurvival during oxidant stress both in the kidney and in cultured mouse proximal tubule (TKPTS) cells and demonstrated concomitant activation of ERK as well as the cyclic adenosine monophosphate (cAMP)-responsive element binding protein (CREB), during survival in vitro. We now show that CREB is a necessary prosurvival target of ERK. METHODS: Ischemia/reperfusion (I/R) injury was induced in 129Sv mice. Oxidant stress was induced by hydrogen peroxide (H(2)O(2)) in TKPTS cells. Activation of CREB was determined by immunohistochemistry and Western blotting. Inhibition and activation of CREB was achieved by mutant or activated CREB-containing adenoviruses in vitro. The effects of oxidant stress on cell survival, CREB binding, and CREB-mediated transcription was determined by cell counting, gelshift analysis, and luciferase assay, respectively. RESULTS: I/R activates CREB in the surviving distal nephron segments of the kidney. Inhibition of ERK and CREB abrogates survival after 0.5 mmol/L H(2)O(2) treatment, while overexpression of CREB ameliorates necrotic death caused by 1 mmol/L H(2)O(2). Inhibition of ERK also inhibited CREB activation. Binding of phosphorylated CREB to a CREB oligonucleotide was significantly increased after 0.5 mmol/L H(2)O(2) but decreased after 1 mmol/L H(2)O(2). Similarly, CREB-mediated transcription was significantly increased after 0.5 mmol/L H(2)O(2) treatment, while 1 mmol/L H(2)O(2) inhibited it. Interestingly, transcription from the CREB-driven bcl-2 promoter was unchanged after 0.5 mmol/L but decreased after 1 mmol/L H(2)O(2) treatment in agreement with Western blot studies. CONCLUSION: We show that survival during oxidant stress is mediated through CREB and identification of its downstream targets will reveal important survival pathways.

Animals↗

PubMed2004

Cisplatin-induced cell death is EGFR/src/ERK signaling dependent in mouse proximal tubule cells.

Cisplatin treatment induces extensive death of the proximal tubules in mice. We also demonstrated that treatment of immortalized mouse proximal tubule cells (TKPTS) with 25 microM cisplatin induces apoptotic death in vitro. Here, we demonstrate that members of the MAPKs such as ERK, JNK, and p38 are all activated after cisplatin treatment both in vivo and in vitro. Because MAPKs mediate cell survival and death, we studied their role in cisplatin-induced cell death in vitro. Apoptosis was confirmed by cell morphology, fluorescence-activated cell-sorting analysis, annexin V/propidium iodide binding, and caspase-3 activation in TKPTS cells. Inhibition of ERK, but not JNK or p38, abolished caspase-3 activation and apoptotic death, suggesting a prodeath role of ERK in cisplatin-induced injury. We also determined that cisplatin-induced ERK as well as caspase-3 activation are epidermal growth factor receptor (EGFR) and c-src dependent because inhibition of these genes inhibited ERK and caspase-3 activation and attenuated apoptotic death. These results suggest that caspase-3 mediates cisplatin-induced cell death in TKPTS cells via an EGFR/src/ERK-dependent pathway. We also suggest that the prodeath effect of ERK is injury type dependent because during oxidant injury, ERK supports survival rather than death in the same cells. We propose that injury-specific outcome diverges downstream from ERK in cisplatin- or H(2)O(2)-mediated cell survival and death.

Animals↗

PubMed2004

Activation of ERK or inhibition of JNK ameliorates H(2)O(2) cytotoxicity in mouse renal proximal tubule cells.

BACKGROUND: Our previous studies suggest that the balance between the activation of extracellular signal-regulated kinase (ERK) and the c-Jun N-terminal/stress-activated protein kinase (JNK) might determine cell fate following oxidant injury in vivo. METHODS: The mouse proximal tubule cell line (TKPTS) was used to study hydrogen peroxide (H(2)O(2))-induced death and survival. The role of ERK and JNK in this process was studied by using adenoviruses that contain either a constitutively active mitogen-activated protein kinase kinase 1 (MEK1) or a dominant-negative JNK. Acridine orange plus ethidium bromide staining was applied to distinguish between viable, apoptotic, and necrotic cells following H(2)O(2) treatment. We analyzed cell cycle events by fluorescence-activated cell sorter (FACS) analysis and the phosphorylation status of ERK and JNK by Western blotting. RESULTS: TKPTS cells survived a moderate level of oxidative stress (0.5 mM/L H(2)O(2)) via temporary growth arrest, while high dose of H(2)O(2) (1 mM/L) caused extensive necrosis. Survival was associated with activation of both ERK and JNK, while death was associated with JNK activation only. Prior adenovirus-mediated up-regulation of ERK or inhibition of JNK function increased the survival (8- or 7-fold, respectively) of TKPTS cells after 1 mmol/L H(2)O(2) treatment. Interestingly, ERK activation and, thus, survival was associated with growth arrest not proliferation. CONCLUSION: We demonstrate that oxidant injury-induced necrosis could be ameliorated by either up-regulation of endogenous ERK or by inhibition of JNK-related pathways. These results directly demonstrate that the intracellular balance between prosurvival and prodeath mitogen-activated protein kinases (MAPKs) determine proximal tubule cell survival from oxidant injury and reveal possible mediators of survival.

Adenoviridae↗

PubMed2003

Differentiation-dependent expression of signal transducers and activators of transcription (STATs) might modify responses to growth factors in the cancers of the head and neck.

Overexpression of the epidermal growth factor receptor (EGFR) in the cancers of the head and neck is well demonstrated. In addition, copy numbers of the EGFR mRNA were significantly higher in poorly differentiated tumors than in tumors that had a differentiated phenotype. Studies by others also showed that the constitutively activated signal transducer and activator of transcription-3 (STAT3), but not STAT1, is required for EGFR-mediated cell growth. Our aim was to reveal if STAT expression is differentiation-dependent and thus, might respond to exogenous stimuli in a differentiation-dependent manner. Both reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry revealed that expression of STAT1 was high in well/moderately differentiated tumors in vivo. In contrast, STAT3 was expressed in poorly differentiated tumors. In vitro experiments showed that differentiated primary oral keratinocytes expressed higher levels of STAT1, but lower levels of STAT3 than did their undifferentiated counterparts. Epidermal growth factor treatment of oral keratinocytes with various degrees of differentiation showed the maximal induction of cyclin D1 in undifferentiated cells. Our findings suggest that the level of differentiation might modulate the outcome of EGFR signaling (i.e. cyclin D1 transcription), due to the differentiation-associated intracellular balance of transcriptional regulators (STAT1 versus STAT3).

Biopsy↗

PubMed2003

A functional interferon regulatory factor-1 (IRF-1)-binding site in the upstream regulatory region (URR) of human papillomavirus type 16.

We recently identified a new enhancer element (HIRE-1, HPV-Interferon Responsive Element-1) in the upstream regulatory region (URR) of human papillomavirus (HPV) type 16. HIRE-1 is located upstream from and in close proximity to the TATA box. HIRE-1 is 1 nt shorter in its 5' sequence in comparison to a consensus IRF-1 binding site (IRF-E). Gel shift analyses clearly demonstrated that HIRE-1 is capable of binding IRF-1 in response to interferon-gamma (IFN-gamma) treatment. In a reporter system, HIRE-1 stimulated transcription in response to IRF-1 or IFN gamma from both a heterologous or the homologous (p97) promoter in a dose-dependent manner. Mutations in the core binding sequence strongly decreased this enhancer activity. Interestingly, HIRE-1 stimulated transcription in the context of the full URR in a cell-type-specific manner, thereby suggesting the role of other cell-type-specific factors that might counteract with its function. Thus, our results may explain the inconsistent clinical and experimental results observed following IFN treatment of cervical lesions or cells. Also, this new enhancer may have an important function during inflammatory responses against HPV type 16.

5' Flanking Region↗

PubMed2003

Cisplatin nephrotoxicity.

Cisplatin remains a major antineoplastic drug for the treatment of solid tumors. Its chief dose-limiting side effect is nephrotoxicity, which evolves slowly and predictably after initial and repeated exposure. The kidney accumulates cisplatin to a higher degree than other organs perhaps via mediated transport. Functionally, reduced renal perfusion and a concentrating defect characterize its nephrotoxicity, whereas morphologically necrosis of the terminal portion of the proximal tubule and apoptosis predominantly in the distal nephron characterize its effects on cell fate. Among the earliest reactions of the kidney to cisplatin is the activation of the MAPK cascade and molecular responses typical of the stress response. Repression of genes characteristic of the mature phenotype of the kidney, especially those serving transport function of the kidney, is also prominent. Metabolic responses, cell cycle events and the inflammatory cascade seem to be important determinants of the degree of renal failure induced by cisplatin. Manipulation of these responses may be exploited to reduce its toxicity clinically.

Antineoplastic Agents↗

PubMed2002

Interleukin-10 induces transcription of the early promoter of human papillomavirus type 16 (HPV16) through the 5'-segment of the upstream regulatory region (URR).

The effects of various proinflammatory cytokines on the transcription of human papillomaviruses (HPVs) have been demonstrated. On the other hand, the role of anti-inflammatory cytokines has not been elaborated, despite the fact that levels of interleukin-10 (IL-10) have been found significantly elevated in cervical dysplasias or carcinomas as well as in the cervix of HIV-positive individuals. These conditions are also associated with elevated viral transcription. Thus, the impact of IL-10 on HPV transcription might be important in pathogenesis of cervical lesions in both immunocompetent or immunosuppressed individuals. In this paper we describe the effects of IL-10 on the transcription of HPV type 16. We found that treatment of HPV 16-positive cervical carcinoma cells with IL-10 increased mRNA levels of the E7 early gene at the level of transcription. Similarly, IL-10 significantly and dose-dependently induced the transcription from the HPV early promoter in a reporter system. Employing deletion mutants we determined that this induction is mapped to the 5' segment of the URR. Transient transfection of an antisense-STAT3-expression vector abolished IL-10-induced reporter activity as well as HPV 16 E7 expression. This suggests that STAT3 either directly binds to the URR and stimulates transcription or affects expression and/or binding of transcription factors that bind to the 5'-region. Our findings suggest a mechanism by which--in addition to its immunosuppressive effects--IL-10 might enhance persistence and progression of HPV-related lesions under conditions (e.g. dysplastic progression, HIV infection) when the cytokine expression in the cervical microenvironment changes.

5' Flanking Region↗

PubMed2002

Suppression of growth by all-trans retinoic acid requires prolonged induction of interferon regulatory factor 1 in cervical squamous carcinoma (SiHa) cells.

All-trans retinoic acid (ATRA) suppresses growth of cervical dysplasias in vivo, although the sensitivity to retinoids is frequently lost during cervical carcinogenesis. It has been suggested that prolonged treatment or use of higher doses of retinoids might offer favorable response rates. We found SiHa cervical squamous carcinoma cells that were virtually resistant to ATRA-induced growth-inhibitory effects at physiological doses (10(-7 to) 10(-6) M) to be more responsive at pharmacological doses (10(-5 to) 10(-4) M). The growth inhibition by high-dose (10(-4) M) ATRA was associated with a sustained activation of interferon regulatory factor 1 (IRF-1), while a low dose (10(-6) M) of ATRA activated IRF-1 only transiently. Antisense IRF-1 inhibited the high-dose (10(-4) M), ATRA-mediated growth arrest; forced expression of IRF-1 caused a significant reduction in cell growth. High-dose (10(-4) M) ATRA increased binding of NF-kappaB and STAT1 proteins to sequences that originated from the IRF-1 promoter region, while low-dose (10(-6) M) ATRA induced only NF-kappaB binding. A delayed tyrosine phosphorylation of the signal transducer and activator of transcription-1 (STAT1) was observed after high-dose (10(-4) M) but not low-dose (10(-6) M) ATRA treatment. In agreement with this, induction of IRF-1 mRNA by ATRA was only modest and transient in a STAT1 knockout cell line, suggesting the importance of STAT1 in sustained IRF-1 expression. Our data showed that ATRA is capable of inducing dose-dependent cellular changes, which might be appropriate to overcome resistance to retinoids that frequently develops during cervical carcinogenesis.

Antineoplastic Agents↗

PubMedDate not supplied

All-trans-retinoic acid activates caspase-1 in a dose-dependent manner in cervical squamous carcinoma cells.

Earlier we observed that all-trans-retinoic acid (ATRA) dose-dependently suppressed the growth of cervical carcinoma cells. Suppression of growth required sustained activation of interferon regulatory factor 1 (IRF-1), which was achieved by high-dose (10(-4) M), but not low-dose (10(-6) M), ATRA treatment. In this paper we examine the role of IRF-1 in cell death that accompanied the growth suppression in high-dose ATRA-treated cells. We found that high-dose, but not low-dose, ATRA treatment activated caspase-1 in those cervical carcinoma cells. Transient transfection of an antisense-IRF-1 construct diminished high-dose ATRA-mediated caspase-1 activation. On the other hand, ATRA was not able to induce caspase-1 expression in a STAT1 (signal transducer and activator of transcription 1) knockout cell line, but transient transfection of STAT1 restored it. These results suggested the importance of both IRF-1 and STAT1 in high-dose ATRA-induced activation of caspase-1. Our results might be useful in the treatment of retinoid-resistant cervical neoplasias.

Antineoplastic Agents↗

PubMedDate not supplied

Dose-dependent activation of p21WAF1 transcription by all-trans-acid in cervical squamous carcinoma cells.

SiHa cervical squamous carcinoma cells were resistant to ATRA-induced growth inhibitory effect at a physiological dose (10(-6) M), but responsive at a pharmacological dose (10(-4) M). The observed growth arrest was associated with increased levels of the interferon regulatory factor-1 (IRF-1). IRF-1 is a known inhibitor of cell growth, and thus our aim was to identify the downstream target of this growth inhibitory function. We found that, as with the induction of IRF-1, levels of p21WAF1 were similarly dose-dependently induced by ATRA. Semiquantitative RT-PCR, Western blotting, gel-shift analysis (EMSA), antisense gene expression and application of a STAT1-knockout cell line demonstrated that activation of the p21WAF1 gene was IRF-1 and STAT1-dependent. We concluded that activation of STAT1 and IRF-1 is crucial for the growth inhibitory action of ATRA, which is associated with the activation of p21WAF1. These results might be useful in chemoprevention of cervical cancers.

Antineoplastic Agents↗

PubMedDate not supplied

All-trans retinoic acid (ATRA) suppresses transcription of human papillomavirus type 16 (HPV16) in a dose-dependent manner.

Earlier we found that SiHa cervical squamous carcinoma cells that harbor HPV type 16 respond to ATRA treatment in a dose-dependent manner: high-dose (10(-5)-10(-4) M) but not low-dose (10(-7)-10(-6) M) ATRA induced growth arrest. Growth of HPV-infected cells is highly dependent on the expression of the viral E6/E7 proteins. Thus, targeting expression of the viral E6/E7 genes might influence growth properties of HPV-infected cells. Here, we demonstrated that high-dose ATRA inhibited expression of HPV16 E7 through suppression of the HPV16 promoter (p97) activity. Gelshift assay (EMSA) revealed that binding of the AP-1 transcription factor to an oligonucleotide originated from the HPV type 16 promoter was diminished after high-dose, but not low-dose ATRA treatment. This suggests that high-dose ATRA suppresses HPV 16 promoter activity, at least in part, via a decreased AP-1 binding. Our data might be useful in treatment of cervical dysplasias and/or carcinomas.

Antineoplastic Agents↗