Hypoxia-inducible factor 1 transcriptional activity in endothelial cells is required for acute phase cardioprotection induced by ischemic preconditioning.

Infarction occurs when myocardial perfusion is interrupted for prolonged periods of time. Short episodes of ischemia and reperfusion protect against tissue injury when the heart is subjected to a subsequent prolonged ischemic episode, a phenomenon known as ischemic preconditioning (IPC). Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that mediates adaptive responses to hypoxia/ischemia and is required for IPC. In this study, we performed a cellular and molecular characterization of the role of HIF-1 in IPC. We analyzed mice with knockout of HIF-1α or HIF-1ß in Tie2(+) lineage cells, which include bone marrow (BM) and vascular endothelial cells, compared with control littermates. Hearts were subjected to 30 min of ischemia and 120 min of reperfusion, either as ex vivo Langendorff preparations or by in situ occlusion of the left anterior descending artery. The IPC stimulus consisted of two cycles of 5-min ischemia and 5-min reperfusion. Mice lacking HIF-1α or HIF-1ß in Tie2(+) lineage cells showed complete absence of protection induced by IPC, whereas significant protection was induced by adenosine infusion. Treatment of mice with a HIF-1 inhibitor (digoxin or acriflavine) 4 h before Langendorff perfusion resulted in loss of IPC, as did administration of acriflavine directly into the perfusate immediately before IPC. We conclude that HIF-1 activity in endothelial cells is required for acute IPC. Expression and dimerization of the HIF-1α and HIF-1ß subunits is required, suggesting that the heterodimer is functioning as a transcriptional activator, despite the acute nature of the response.

Biobanking of derivatives from radical retropubic and robot-assisted laparoscopic prostatectomy tissues as part of the prostate cancer biorepository network.

BACKGROUND: The goal of the Prostate Cancer Biorepository Network (PCBN) is to develop a biorepository with high-quality, well-annotated specimens obtained in a systematic, reproducible fashion using optimized and standardized protocols, and an infrastructure to facilitate the growth of the resource and its wide usage by the prostate cancer research community. An emerging area of concern in the field of prostate cancer biobanking is an apparent shift in the proportion of surgical procedures performed for prostate cancer treatment from radical retropubic prostatectomy (RRP) to robot-assisted laparoscopic prostatectomy (RALP). Our study aimed to determine the potential impact of the RALP procedure on the detection of known prostate cancer biomarkers, and the subsequent suitability of RALP-derived specimens for prostate cancer biomarker studies. METHODS: DNA and RNA were extracted from RRP and RALP specimens. Quality assessment was conducted using spectrophotometric analysis and RNA was analyzed for RNA integrity number (RIN) and by real-time reverse-transcription PCR (qRT-PCR) for racemase, hepsin, ERG, TMPRSS2-ERG gene fusions, and the microRNAs miR-26a, miR-26b, miR-141, and miR-221. RESULTS: We demonstrate that extraction of derivatives from frozen tissues from RRP and RALP specimens yields samples of equally high quality as assessed by spectrophotometric and RIN analysis. Likewise, expression levels of genes analyzed by qRT-PCR did not differ between RRP and RALP-derived tissues. CONCLUSIONS: Our studies indicate that samples obtained from RALP specimens may be suitable for prostate cancer biomarker studies-an important finding given the current shift in surgical procedures for prostate cancer treatment.

Taxane-induced blockade to nuclear accumulation of the androgen receptor predicts clinical responses in metastatic prostate cancer.

Prostate cancer progression requires active androgen receptor (AR) signaling which occurs following translocation of AR from the cytoplasm to the nucleus. Chemotherapy with taxanes improves survival in patients with castrate resistant prostate cancer (CRPC). Taxanes induce microtubule stabilization, mitotic arrest, and apoptotic cell death, but recent data suggest that taxanes can also affect AR signaling. Here, we report that taxanes inhibit ligand-induced AR nuclear translocation and downstream transcriptional activation of AR target genes such as prostate-specific antigen. AR nuclear translocation was not inhibited in cells with acquired ß-tubulin mutations that prevent taxane-induced microtubule stabilization, confirming a role for microtubules in AR trafficking. Upon ligand activation, AR associated with the minus-end-microtubule motor dynein, thereby trafficking on microtubules to translocate to the nucleus. Analysis of circulating tumor cells (CTC) isolated from the peripheral blood of CRPC patients receiving taxane chemotherapy revealed a significant correlation between AR cytoplasmic sequestration and clinical response to therapy. These results indicate that taxanes act in CRPC patients at least in part by inhibiting AR nuclear transport and signaling. Further, they suggest that monitoring AR subcellular localization in the CTCs of CRPC patients might predict clinical responses to taxane chemotherapy.

Carbohydrate restriction, prostate cancer growth, and the insulin-like growth factor axis.

BACKGROUND: Recent evidence suggests carbohydrate intake may influence prostate cancer biology. We tested whether a no-carbohydrate ketogenic diet (NCKD) would delay prostate cancer growth relative to Western and low-fat diets in a xenograft model. METHODS: Seventy-five male SCID mice were fed a NCKD (84% fat-0% carbohydrate-16% protein kcal), low-fat (12% fat-72% carbohydrate-16% protein kcal), or Western diet (40% fat-44% carbohydrate-16% protein kcal). Low-fat mice were fed ad libitum and the other arms fed via a modified-paired feeding protocol. After 24 days, all mice were injected with LAPC-4 cells and sacrificed when tumors approached 1,000 mm(3). RESULTS: Despite consuming equal calories, NCKD-fed mice lost weight (up to 15% body weight) relative to low-fat and Western diet-fed mice and required additional kcal to equalize body weight. Fifty-one days after injection, NCKD mice tumor volumes were 33% smaller than Western mice (rank-sum, P = 0.009). There were no differences in tumor volume between low-fat and NCKD mice. Dietary treatment was significantly associated with survival (log-rank, P = 0.006), with the longest survival among the NCKD mice, followed by the low-fat mice. Serum IGFBP-3 was highest and IGF-1:IGFBP-3 ratio was lowest among NCKD mice while serum insulin and IGF-1 levels were highest in Western mice. NCKD mice had significantly decreased hepatic fatty infiltration relative to the other arms. CONCLUSIONS: In this xenograft model, despite consuming more calories, NCKD-fed mice had significantly reduced tumor growth and prolonged survival relative to Western mice and was associated with favorable changes in serum insulin and IGF axis hormones relative to low-fat or Western diet.

The l2 minor capsid protein of human papillomavirus type 16 interacts with a network of nuclear import receptors.

The L2 minor capsid proteins enter the nucleus twice during viral infection: in the initial phase after virion disassembly and in the productive phase when, together with the L1 major capsid proteins, they assemble the replicated viral DNA into virions. In this study we investigated the interactions between the L2 protein of high-risk human papillomavirus type 16 (HPV16) and nuclear import receptors. We discovered that HPV16 L2 interacts directly with both Kapbeta(2) and Kapbeta(3). Moreover, binding of Ran-GTP to either Kapbeta(2) or Kapbeta(3) inhibits its interaction with L2, suggesting that the Kapbeta/L2 complex is import competent. In addition, we found that L2 forms a complex with the Kapalpha(2)beta(1) heterodimer via interaction with the Kapalpha(2) adapter. In agreement with the binding data, nuclear import of L2 in digitonin-permeabilized cells could be mediated by either Kapalpha(2)beta(1) heterodimers, Kapbeta(2), or Kapbeta(3). Mapping studies revealed that HPV16 L2 contains two nuclear localization signals (NLSs), in the N terminus (nNLS) and C terminus (cNLS), that could mediate its nuclear import. Together the data suggest that HPV16 L2 interacts via its NLSs with a network of karyopherins and can enter the nucleus via several import pathways mediated by Kapalpha(2)beta(1) heterodimers, Kapbeta(2), and Kapbeta(3).