Additional file 1 of Diagnostic accuracy of procalcitonin in adult non-neutropenic cancer patients with suspected infection: a systematic review and meta-analysis

Appendix

TPT-induced elevation of MHC I and IFN-β is DNA replication-, but not transcription-, dependent.

<p>(<b>A</b>) TPT-induced MHC I expression require active DNA synthesis. ZR-75-1 cells were pre-treated with APH (10 µM) for 30 min, followed by 0.1% DMSO or TPT (40 nM) treatment for 1 hr, and subsequent incubation in drug-free medium for 4 days. Expression of MHC I was then measured by immunoblotting. (<b>B</b>) TPT-induced MHC I expression is independent of transcription. ZR-75-1 cells were pre-treated with the transcription inhibitor DRB (150 µM) for 30 min, and then treated with TPT as described in (<b>A</b>). Expression of MHC I was measured by immunoblotting. (<b>C</b>) Conditioned medium-induced MHC I expression in recipient cells requires active DNA synthesis in TPT-treated donor cells. Recipient ZR-75-1 cells were incubated with conditioned media from TPT-treated donor cells as described in (<b>A</b>). The total expression of cellular MHC I in recipient cells was then measured by immunoblotting. (<b>D</b>) TPT-induced IFN-β mRNA expression requires active DNA synthesis. ZR-75-1 cells were pretreated with APH (10 µM) or DRB (150 µM) for 30 min prior to co-incubation with TPT (40 nM, 1 hr), followed by drug-free incubation for 3 days. Total RNAs were isolated for real-time RT-PCR analysis. The experiments have been repeated three times. *: <i>p</i>-value<0.05. The error bar indicates standard deviation.</p

Cancer chemotherapeutics and ionizing radiation induce MHC I expression in breast cancer cells.

<p>(<b>A</b>) Cancer chemotherapeutics induce elevated expression of total cellular MHC I in breast cancer ZR-75-1 cells. Cells were treated with topotecan (40 nM), etoposide (1 µM), cisplatin (6 µM), paclitaxel (3 µM), and vinblastine (6 nM) for 24 hrs, followed by incubation in drug-free medium for 3 days. Expression of total cellular MHC I was measured by immunoblotting. (<b>B</b>) Cancer chemotherapeutics induce cell-surface expression of MHC I in ZR-75-1 cells. ZR-75-1 cells were treated with different anticancer drugs as described in (<b>A</b>). Expression of cell-surface MHC I was then determined by FACS analysis. (<b>C</b>) Conditioned media from γ-irradiated cells induce elevated total cellular expression of MHC I in radiation-naïve recipient cells. Donor (ZR-75-1) cells were irradiated with γ-ray (2 Gy) for 3, 4, and 5 times. The conditioned media were then transferred to radiation-naïve recipient (ZR-75-1) cells. After 48 hrs, cell lysates were harvested for immunoblotting analysis. (<b>D</b>) Induction of MHC I expression in drug naïve recipient cells depends on IFN-β in the conditioned medium from irradiated donor cells. Radiation-naïve recipient (ZR-75-1) cells were incubated with the conditioned medium from irradiated (2 Gy×4) donor (ZR-75-1) cells in the presence of the neutralizing antibody against IFN-α (anti-α) or IFN-β (anti-β) for two days. Cell-surface MHC I in the recipient (ZR-75-1) cells was then measured by FACS analysis.</p

Increased secretion of IFN-β is responsible for TPT-induced MHC I expression.

<p>(<b>A</b>) Conditioned media from TPT-treated cells stimulate total cellular expression of MHC I in drug-naïve cells. ZR-75-1 cells were treated with 40 nM TPT for 1 hr, followed by incubation in drug-free medium for 4 days. Drug-naïve ZR-75-1 cells were then replenished with the conditioned media for another 2-day incubation. The total expression of cellular MHC I in recipient cells was measured by immunoblotting. (<b>B</b>) Conditioned media from TPT-treated cells stimulate cell-surface expression of MHC I in drug-naïve recipient cells. Recipient ZR-75-1 cells were treated with the conditioned media as described above, and the cell-surface MHC I expression was measured by FACS analysis. (<b>C</b>) TPT treatment increases mRNA levels of various cytokines in ZR-75-1 cells. Cells were treated with TPT (40 nM) for 1 hr, followed by incubation in drug-free medium for 3 days. Total RNAs were harvested for analysis by real-time RT-PCR. The experiments have been repeated twice. *: <i>p</i>-value<0.05. The error bar indicates standard deviation. (<b>D</b>) Purified IFN-β induces total cellular expression of MHC I. ZR-75-1 cells were treated with recombinant human IFN-β (500 U/ml) for 2 days. The total expression of cellular MHC I in recipient cells was measured by immunoblotting.</p

Cancer chemotherapeutics induce IFN-β secretion (ELISA assay).

<p>ZR-75-1 cells were treated with topotecan (40 nM), etoposide (1 µM), cisplatin (6 µM), paclitaxel (3 µM), or vinblastine (6 nM) for 24 hrs, aspirated and washed, followed by incubation in drug-free medium for 3 days. Levels of IFN-β protein in the culture media were then measured by ELISA.</p

TPT-induced MHC I expression requires NF-κB activation, but not apoptotic caspases.

<p>(<b>A</b>) The NF-κB inhibitor BAY 11-7085 (BAY) blocks TPT-induced MHC I expression. ZR-75-1 cells were pretreated with BAY (10 µM) for 30 min, followed by co-incubation with 0.1% DMSO or TPT (40 nM) for 1 hr, and subsequent drug-free incubation for 4 days. Total cellular MHC I was then analyzed by immunoblotting. (<b>B</b>) The conditioned medium-induced MHC I in recipient cells requires NF-κB activation in TPT-treated donor cells. Drug- naïve recipient ZR-75-1 cells were incubated with the conditioned medium from TPT-treated donor cells as described in (<b>A</b>). Expression of total cellular MHC I in recipient cells was then analyzed by immunoblotting. (<b>C</b>) TPT-induced IFN-β mRNA expression requires NF-κB activation. ZR-75-1 cells were pretreated with BAY (10 µM) for 30 min prior to co-incubation with TPT (40 nM, 1 hr), followed by drug-free incubation for 3 days. Total RNAs were isolated for real-time RT-PCR analysis. The experiments have been repeated three times. *: <i>p</i>-value <0.05. The error bar indicates standard deviation. (<b>D</b>) BAY specifically blocks IκBα degradation induced by TNF-α treatment. ZR-75-1 cells were treated with TNF-α (10 ng/ml) for 10 min, in the presence or absence of BAY (10 µM). Expression of IκBα was measured by immunoblotting. (<b>E</b>) TPT-induced MHC I expression is independent of caspase activation. ZR-75-1 cells were pretreated with the pan-caspase inhibitor Z-VAD-FMK (Z-VAD, 10 µM) for 1 hr prior to co-incubation with TPT (40 nM, 1 hr), followed by continued incubation in drug-free medium for 4 days. Cell lysates were then immunoblotted with the anti-MHC I antibody. (<b>F</b>) Staurosporine-induced PAPR-1 cleavage requires caspase activation. ZR-75-1 cells were pretreated with Z-VAD-FMK (10 µM) for 1 hr followed by co-incubation with staurosporine (STS, 0.5 µM) for 6 hrs. PARP-1 cleavage was then measured by immunoblotting.</p

TPT induces elevated expression of both total and cell-surface MHC I in breast cancer cells.

<p>(<b>A</b>) TPT induces elevated expression of total cellular MHC I in breast cancer cells. Breast cancer cell lines ZR-75-1, T47D, MDA-MB-231 (MB-231) and MCF-7 were treated with TPT (40, 200, 20, and 40 nM, respectively) for 4 days. Expression of total cellular MHC I was measured by immunoblotting using a polyclonal antibody against MHC I. (<b>B</b>) The concentration effect of TPT on total cellular expression of MHC I. ZR-75-1 cells were treated with increasing concentrations of topotecan (10 nM to 1 µM) for 4 days, followed by immunoblotting for total cellular MHC I expression. (<b>C</b>) Acute (1 hr) TPT exposure stimulates total cellular expression of MHC I. ZR-75-1 cells were treated with TPT (40 nM) for increasing durations of time (0, 1, 6 and 24 hrs), followed by incubation in drug-free medium for a total combined incubation of 4 days. Total cellular expression of MHC I was measured by immunoblotting. (<b>D</b>) TPT induces elevated cell-surface expression of MHC I. ZR-75-1 cells were treated with 40 nM of TPT for 1 hr, followed by incubation in drug-free medium for 4 days. Cell-surface MHC I was measured by FACS analysis using the pan-MHC I monoclonal antibody, W6/32.</p

IFN-β signaling through type I IFN receptor is responsible for the TPT-induced MHC I expression.

<p>(<b>A</b>) IFNAR1 knockdown abolishes TPT-induced total cellular MHC I expression. On the left panel, ZR-75-1 cells were mock-transfected, or transfected with control siRNA or IFNAR1-specific siRNA. 48 hrs post-transfection, cells were treated with TPT (or 0.1% DMSO as control) for 1 hr, followed by incubation in drug-free medium for 4 days. Lysates were analyzed by immunoblotting. On the right panel, ZR-75-1 cells were transfected with control or IFNAR1-specific siRNA. 72 hrs post-transfection, cells were harvested and immunoblotted with an anti-IFNAR1 antibody. (<b>B</b>) Neutralizing antibody against IFN-β, but not IFN-α, blocks the TPT-induced cell-surface MHC I expression. ZR-75-1 cells were treated with TPT (40 nM or 0.1% DMSO as control) for 1 hr, followed by incubation in drug-free medium with or without neutralizing antibody against IFN-α (anti-α, 1.44×10³ NU/ml) or IFN-β (anti-β, 296 NU/ml). Cell-surface MHC I expression was measured by FACS analysis. (<b>C</b>) Neutralizing antibody against IFN-β, but not IFN-α, blocks the TPT-induced ISG15. Antibody and TPT treatments of ZR-75-1 cells were performed exactly the same as described in (<b>B</b>). Lysates were immunoblotted for ISG15 expression. (<b>D</b>) Neutralizing antibody against IFN-α inhibits IFN-α-induced ISG15 expression. ZR-75-1 cells were co-incubated with IFN-α (100 U/ml) in the presence or absence of anti-IFN-α antibody (1.44×10³ NU/ml) for 2 days. Cells were then harvested and immunoblotted with an anti-ISG15 antibody.</p

Identification of the effectors for ISG15 induction in the Ras-dependent signaling pathway.

<p>A, B. siRNA knock-down of interferon-β and the type I IFN receptor reduces ISG15 induction in the Ras-dependent signaling pathway. The oncogenic Ras-transformed MCF-10A cells (10A/H-Ras and 10A/K-Ras) were transiently transfected with control, IFN-β or IFNAR1 siRNA. Cell lysates were collected 72 hrs post-transfection and immunoblotted with anti-ISG15 and anti-IFNAR1 antibodies. C. The PI3K inhibitor reduces ISG15 expression in oncogenic Ras-transformed cells. 10A/H-Ras and 10A/K-Ras cells were treated with 20 µM LY294002, a PI3K inhibitor, for 3 days. Lysates with equal protein amount were immunoblotted with anti-ISG15 antibodies (L: LY294002). D. The NF-κB inhibitor reduces ISG15 expression in oncogenic Ras-transformed cells. 10A/H-Ras and 10A/K-Ras cells were seeded and cultured overnight. Cells were then treated with or without 40 µM Bay 11-7085, an NF-κB inhibitor, for 24 hrs. Cell lysates were analyzed by immunoblotting using anti-ISG15, phospho-IκBα, and IκBα antibodies.</p

X-ray Absorption Spectroscopic Study on Interfacial Electronic Properties of FeOOH/Reduced Graphene Oxide for Asymmetric Supercapacitors

[[abstract]]The effects of growth time and interface between the iron oxyhydroxide (FeOOH) and carbon materials (carbon nanotubes (CNT) and reduced graphene oxide (RGO)) to form an asymmetric supercapacitor was studied by X-ray absorption spectroscopy (XAS) and electrochemical measurements. FeOOH/CNT (FCNT) and FeOOH/RGO (FRGO) were successfully synthesized by a simple spontaneous redox reaction with FeCl3. The RGO functions as an ideal substrate, providing rich growth sites for FeOOH, and it is believed to facilitate the transport of electrons/ions across the electrode/electrolyte interface. FRGO has been identified as a supercapacitor and found to exhibit significantly greater capacitance than FCNT. To gain further insight into the effects of growth times and the interface of FeOOH for FCNT and FRGO, the electronic structures of FCNT and FRGO with various FeOOH growth times were elucidated by XAS. The difference between the surface electronic structures of CNT and RGO yields different nucleation and growth rates of FeOOH of FeOOH. RGO with excellent interface properties arises from a high degree of covalent functionalization, and/or defects make it favorable for FeOOH growth. FRGO is therefore a promising electrode material for use in the fabrication of asymmetric supercapacitors. In this work, coupled XAS and electrochemical measurements reveal the electronic structure of the interface between FeOOH and the carbon materials and the capacitance performance of asymmetric supercapacitors, which are very useful in the fields of nanomaterials and nanotechnology, especially for their applications in storing energy[[notice]]補正完