Caspase protocols in mice.

Members of the caspase family of proteases are evolutionarily conserved cysteine proteases that play a crucial role as the central executioners of the apoptotic pathway. Since the discovery of caspases, many methods have been developed to detect their activation and are widely used in basic and clinical studies. In a mouse tissue, caspase activation can be monitored by cleavage of caspase-specific synthetic substrates and by detecting cleaved caspase by western blot analysis of the tissue extract.

Meprin A and meprin alpha generate biologically functional IL-1beta from pro-IL-1beta.

The present study demonstrates that both oligomeric metalloendopeptidase meprin A purified from kidney cortex and recombinant meprin alpha are capable of generating biologically active IL-1beta from its precursor pro-IL-1beta. Amino-acid sequencing analysis reveals that meprin A and meprin alpha cleave pro-IL-1beta at the His(115)-Asp(116) bond, which is one amino acid N-terminal to the caspase-1 cleavage site and five amino acids C-terminal to the meprin beta site. The biological activity of the pro-IL-1beta cleaved product produced by meprin A, determined by proliferative response of helper T-cells, was 3-fold higher to that of the IL-1beta product produced by meprin beta or caspase-1.

Autophagy delays apoptosis in renal tubular epithelial cells in cisplatin cytotoxicity.

One of the major side effects of cisplatin chemotherapy is toxic acute kidney injury due to preferential accumulation of cisplatin in renal proximal tubule epithelial cells and the subsequent injury to these cells. Apoptosis is known as a major mechanism of cisplatin-induced cell death in renal tubular cells. We have also recently demonstrated that autophagy induction is an immediate response of renal tubular epithelial cell exposure to cisplatin.

Autophagy is associated with apoptosis in cisplatin injury to renal tubular epithelial cells.

Autophagy has emerged as another major "programmed" mechanism to control life and death much like "programmed cell death" is for apoptosis in eukaryotes. We examined the expression of autophagic proteins and formation of autophagosomes during progression of cisplatin injury to renal tubular epithelial cells (RTEC). Autophagy was detected as early as 2-4 h after cisplatin exposure as indicated by induction of LC3-I, conversion of LC3-I to LC3-II protein, and upregulation of Beclin 1 and Atg5, essential markers of autophagy.

Expression of tissue factor in prostate cancer correlates with malignant phenotype.

Tissue factor (TF), apart from its established role in hemostasis, has been implicated in promoting angiogenesis and metastasis in a wide array of tumors including prostate cancer. Expression of TF was evaluated in freshly-resected prostate specimens obtained from patients with localized (n=9) and androgen ablated (n=6) disease using real-time reverse transcription-polymerase chain reaction and Western blot analysis. TF was detected in all specimens in both stages of the disease.

Isolated thrombocytopenia induced by thalidomide in a patient with multiple myeloma: case report and review of literature.

Thalidomide is being increasingly used in hematology and oncology. Its use is associated with neuropathy, sedation, edema, fatigue, constipation, and deep venous thrombosis. Cytopenias are unusual, but there are case reports.

Mcl-1 is downregulated in cisplatin-induced apoptosis, and proteasome inhibitors restore Mcl-1 and promote survival in renal tubular epithelial cells.

Mcl-1 is an antiapoptotic member of the Bcl-2 family that plays an important role in cell survival. We demonstrate that proteasome-dependent regulation of Mcl-1 plays a critical role in renal tubular epithelial cell injury from cisplatin. Protein levels of Mcl-1 rapidly declined in a time-dependent manner following cisplatin treatment of LLC-PK(1) cells.

Thrombin receptor expression is upregulated in prostate cancer.

Background: Aberrant expression of protease-activated receptors (PARs) has been associated with increased angiogenesis, tumor growth, and metastasis of various cancers. We assessed the status of PAR1 expression in prostate cancer.

Methods: The study compared the abundance levels of PAR1 RNA and protein using real-time reverse-transcriptase polymerase chain reaction and immunoblotting in freshly resected prostate tissues from early localized-stage disease (n=9) to those from patients with advanced metastatic disease (n=7).

p53-dependent caspase-2 activation in mitochondrial release of apoptosis-inducing factor and its role in renal tubular epithelial cell injury.

We demonstrate the role of p53-mediated caspase-2 activation in the mitochondrial release of apoptosis-inducing factor (AIF) in cisplatin-treated renal tubular epithelial cells. Gene silencing of AIF with its small interfering RNA (siRNA) suppressed cisplatin-induced AIF expression and provided a marked protection against cell death. Subcellular fractionation and immunofluorescence studies revealed cisplatin-induced translocation of AIF from the mitochondria to the nuclei.

Differential toxicity of anthracyclines on cultured endothelial cells.

Anthracyclines are known for their endothelial toxicity. Newer derivatives may have fewer toxic effects on endothelium. The authors therefore evaluated the effects of doxorubicin, doxorubicin analogs (daunorubicin, idarubicin), and pegylated liposomal doxorubicin (doxil) in human coronary artery endothelial cells (HCAECs).

Stage-specific characterization of the vascular endothelial growth factor axis in prostate cancer: expression of lymphangiogenic markers is associated with advanced-stage disease.

Purpose: The vascular endothelial growth factor (VEGF) family plays a critical role in tumor angiogenesis and lymphangiogenesis. We characterized, at the mRNA and protein levels, the expression of VEGF-A and VEGF-D and their cognate receptors, VEGFR-1, VEGFR-2, and VEGFR-3 in early- and advanced-stage prostate cancer specimens.

Experimental Design: The levels of VEGF-A and VEGF-D mRNA in early- and advanced-stage specimens were compared using an angiogenic gene array and were confirmed by quantitative real-time PCR.

Regulation of caspase-3 and -9 activation in oxidant stress to RTE by forkhead transcription factors, Bcl-2 proteins, and MAP kinases.

Cytotoxicity to renal tubular epithelial cells (RTE) is dependent on the relative response of cell survival and cell death signals triggered by the injury. Forkhead transcription factors, Bcl-2 family member Bad, and mitogen-activated protein kinases are regulated by phosphorylation that plays crucial roles in determining cell fate. We examined the role of phosphorylation of these proteins in regulation of H(2)O(2)-induced caspase activation in RTE.

Role of coagulation and fibrinolytic system in prostate cancer.

Patients with prostate carcinoma paradoxically have both a hypercoagulable state and a bleeding diathesis. Hypercoagulability manifested by venous and arterial thrombosis has been documented in several large clinical trials. However, many investigators have reported a high risk of postoperative bleeding in prostate cancer patients.

Potential anticancer effects of statins: fact or fiction?

Deregulation of any of the steps in cell growth, proliferation and apoptosis may result in its malignant transformation. Statins, along with their lipid-lowering potential, modify several processes in the cell cycle. These agents inhibit cell proliferation and arrest cell cycle progression by interrupting growth-promoting signals.

Cloning and expression of rat caspase-6 and its localization in renal ischemia/reperfusion injury.

Background: Caspase-6 is an important member of the executioner caspases in the caspase family of cell death proteases. The executioner caspases are the major active caspases detected in apoptotic cells and are generally considered to mediate the execution of apoptosis by cleaving and inactivating intracellular proteins. However, the complete characterization of mRNA and protein of caspase-6 in rat and its expression in normal kidney and in disease state has not been previously elucidated.