Role of human copper transporter Ctr1 in the transport of platinum-based antitumor agents in cisplatin-sensitive and cisplatin-resistant cells

Recent studies have shown that the mammalian high-affinity copper transporter encoded by Ctr1 is involved in the uptake of cisplatin. However, the roles of hCtr1 in cisplatin-sensitive and cisplatin-resistant mammalian cells have not been investigated. Here, we show that, of five cisplatin-resistant cell lines, only one (SR2) exhibited substantial reduction in hCtr1 expression as compared with that in its sensitive line small cell lung cancers (SCLC), whereas copper efflux transporters ATP7A and ATP7B were not significantly altered. SR2 exhibited cross-resistance to carboplatin but not to oxaliplatin. Transfection of expression hemagglutinin-tagged hCtr1 cDNA into SCLC and SR2 cells enhanced the uptake of copper, cisplatin, carboplatin, and oxaliplatin, suggesting that hCtr1 transporter can transport these platinum-based drugs. Whereas increased sensitivities to all these platinum drugs were observed in hCtr1-transfected SCLC cells, increased sensitivities to cisplatin and carboplatin but not to oxaliplatin were observed in hCtr1-transfected SR2 cells. These results suggest that SR2 acquired an additional unique intracellular resistance mechanism to oxaliplatin. Finally, using hCtr1 deletion mutants, we showed that the NH2-terminal domain of hCtr1 was involved in transporting all these platinum-based antitumor agents. These results collectively show the importance of hCtr1 in the transport of platinum-based antitumor agents in cisplatin-sensitive and cisplatin-resistant variants.     

Diagnostic accuracy of lung ultrasonography combined with procalcitonin for the diagnosis of pneumonia: a pilot study

Background

The diagnostic value of lung ultrasonography (LUS) and procalcitonin (PCT) in the diagnosis of lung infections is known. No studies evaluated the combination of LUS and PCT for the diagnosis of pneumonia in the emergency department (ED). We evaluated the diagnostic accuracy of the combination of LUS and PCT in the diagnosis of pneumonia.

Methods

Patients with respiratory symptoms of unexplained origin who underwent a chest CT in ED were included in the study if PCT assay was available. LUS was performed before CT and was targeted to the detection of lung consolidations with the morphologic features of pneumonia. A PCT assay was performed at presentation, and cut-off of 0.25 and of 0.5 ng/ml were used to rule-out and rule-in pneumonia. The final diagnosis of pneumonia was established by independent clinicians, on the basis of clinical chart review including CT results.

Results

We enrolled 128 patients and pneumonia was the final diagnosis in 61 (47.7%). In 38 patients (29.7%) LUS and PCT were negative (PCT < 0.25 ng/ml). The overall accuracy, sensitivity and negative predictive value of LUS/PCT were 88.8, 96.7 and 94.7% respectively. Sensitivity of the LUS/PCT test was significantly superior to LUS alone (85.2%) and PCT alone (73.8%) (p < 0.05 for both). Specificity and positive predictive value of the combination of positivity of LUS/PCT (PCT > 0.5 ng/ml) were 94% and 83.3% respectively. Specificity of LUS/PCT was not significantly different to LUS alone (88.1%) (p = 0.125).

Conclusions

The sensitivity of the combination of LUS with PCT for the diagnosis of pneumonia was significantly superior when compared with the sensitivity of LUS and PCT alone.

     

Chronic lithium administration ameliorates 2,4,6-trinitrobenzene sulfonic acid-induced colitis in rats; potential role for adenosine triphosphate sensitive potassium channels

Background and Aim: Inflammatory bowel disease (IBD) is a multi‐factorial disease with an unknown etiology characterized by oxidative stress, leukocyte infiltration and a rise in inflammatory cytokines. This study was conducted to investigate lithium in 2,4,6‐trinitrobenzene sulfonic acid (TNBS)‐induced chronic model of experimental IBD, and the contribution of potassium channels as a possible underlying mechanism. Methods: Experimental IBD was induced in rats by a single colonic administration of 10 mg of TNBS. Lithium, Glibenclamide (a potassium channel blocker), Lithium + Glibenclamide, Cromakalim or Lithium + Glibenclamide + Cromakalim were given twice daily for 7 successive days. At the end of the experiment, macroscopic and histopathologic scores, colonic malondialdehyde (MDA), tumor necrosis factor‐α (TNF‐α) level, and myeloperoxidase (MPO) activity as well as plasma lithium level were assessed. Results: Both macroscopic and histological features of colonic injury were markedly ameliorated by lithium. Likewise, the elevated amounts of MPO and MDA were diminished as well as those of TNF‐α (P < 0.05). Glibenclamide reversed the effect of lithium on these markers, Addition of cromakalim abrogated the effects mediated by glibenclamide and markedly decreased MPO activity, MDA level and TNF‐α content (P < 0.0.05). Macroscopic and microscopic scores and biochemical markers were significantly decreased in Cromakalim‐treated animals. No significant difference was observed between TNBS and Glibenclamide groups. Conclusion: Lithium exerts prominent anti‐inflammatory effects on TNBS‐induced colitis in rats. Potassium channels contribute to these beneficial properties.     

HLA DR15 antigen status does not impact graft-versus-host disease or survival in HLA-matched sibling transplantation for hematologic malignancies

The HLA class II DRB1 antigen DR15 is an important prognostic marker in immune-mediated marrow failure states. DR15 has also been associated with favorable outcomes (reduced acute graft-versus-host disease [aGVHD] and relapse) after allogeneic hematopoietic cell transplant. To elucidate the impact of DR15 on transplantation outcomes, we conducted a retrospective study of 2891 recipients of first allogeneic stem cell transplant from HLA-matched sibling donors for the treatment of acute leukemia, chronic myeloid leukemia, or myelodysplastic syndrome (MDS) between 1990 and 2007. All patients received conventional myeloablative conditioning, T-replete grafts, and cyclosporine plus methotrexate-based GVHD prophylaxis. DNA-based HLA typing allowed categorization of 732 patients (25.3%) as positive and 2159 patients (74.7%) as negative for DRB1*15:01 or *15:02 (DR15). There were no significant differences in baseline characteristics between the HLA DR15 positive and negative groups. In univariate analysis, HLA-DR15 status had no impact on neutrophil engraftment, aGVHD, chronic GVHD (cGVHD), treatment-related mortality, relapse, disease-free survival, or overall survival (OS). In multivariate analysis, DR15 status showed no significant difference in aGVHD, cGVHD, OS, or relapse. In conclusion, DR15 status had no impact on major HLA-matched sibling donor hematopoietic cell transplant outcomes in this large and homogenous cohort of patients with leukemia and MDS.     

D469del-COMP retention in chondrocytes stimulates caspase-independent necroptosis

Mutations in the cartilage oligomeric matrix protein gene (COMP) cause pseudoachondroplasia (PSACH). This dysplasia results from the intracellular retention of mutant COMP protein and premature death of growth-plate chondrocytes. Toward better understanding of these underlying mechanisms, we examined D469del-COMP activation of the unfolded protein response and cell death pathways in rat chondrosarcoma cells. Using an inducible expression system, we examined the effects of D469del-COMP retention after 4 days of mRNA expression and then 5 days without inducing agent. Retention of D469del-COMP stimulated Chop (Ddit3) and Gadd34 (Ppp1r15a) and triggered reactivation of protein translation that exacerbated intracellular retention. High levels of Nox4 and endoplasmic reticulum receptor stress-inducible Ero1β generated reactive oxygen species, causing oxidative stress. Increased expression of Gadd genes and presence of γH2AX indicated that DNA damage was occurring. The presence of cleaved apoptosis inducing factor (tAIF) and the absence of activated caspases indicated that retention of D469del-COMP triggers cell death in chondrocytes by necroptosis, a caspase-independent programmed necrosis. Loss of growth-plate chondrocytes by necroptosis was also found in our pseudoachondroplasia mouse model. These results suggest a model in which D469del-COMP expression induces persistent endoplasmic reticulum stress, oxidative stress, and DNA damage, thus priming chondrocytes for necroptosis. We define for the first time the precise mechanisms underlying D469del-COMP pathology in pseudoachondroplasia and suggest that oxidative stress and AIF may be promising therapeutic targets.     

Multi Drug Resistance-1 (MDR1) Expression in Response to Chronic Diazinon Exposure: An In vitro Study on Caco-2 Cells

To evaluate the effect of chronic exposure to diazinon in the gastrointestinal tract, Caco-2 cells were made resistant by growing in low concentrations of diazinon (0.02 μM) that was gradually increased to 20 μM within 4.5 months. Resistant cells showed significant higher growth in the presence of 15, 45 and 135 μM of diazinon (96.08, 81.80 and 65.16% of control) compared to parent cells (79.71, 71.76 and 29.50% of control, respectively; p < 0.05). P-glycoprotein (P-gp) expression increased significantly in resistant cells (P-gp to beta-actin ratio 0.586 for parent and 1.255 for resistant cells, respectively; p < 0.05) without any alteration in MDR-1 mRNA level (p > 0.05).     

人肺腺癌染色体8p21～p22杂合性缺失精细作图

目的　在染色体 8p2 1 8p2 2界定发生于中国人肺腺癌的杂合性缺失 (LOH)的最小区域 ,为定位克隆肺腺癌相关抑癌基因提供线索。方法　利用 32例肺腺癌患者肿瘤组织检测位于 8p2 1～p2 2的 17个微卫星多态性标记的LOH频率 ,并且探讨各位点LOH与病理分级和临床分期的关系。结果　在 32例肺腺癌患者组织中有 31例 (96 6 7% )存在至少 1个位点的LOH ;主要集中于 3个区域 :位于 8p2 2的D8S2 5 4～ 2 6 1、D8S182 7～ 1731以及D8S1135。所检测位点中仅D8S2 6 1位点的LOH发生频率与肺腺癌分期呈正相关 (P <0 0 5 )。结论　 8p2 2区域可能存在位于D8S2 5 4～ 2 6 1,D8S1135和D8S182 7～ 1731的、与中国人肺腺癌相关的抑癌基因