Assessment value of interleukin-6, procalcitonin, and C-reactive protein early kinetics for initial antibiotic efficacy in patients with febrile neutropenia: A prospective study.

BACKGROUND: This study aims to investigate the early kinetics of interleukin 6 (IL-6), procalcitonin (PCT), and C-reactive protein (CRP) on initial antibiotic efficacy in hematological disorder patients with febrile neutropenia (FN). METHODS: A total of 40 patients with 43 episodes of FN were enrolled and divided into initial antibiotic effective group (IAE group, n = 24) and initial antibiotic ineffective group (IAI group, n = 19). The levels of IL-6, PCT, and CRP before antibacterial treatment (T0), and 12 h (T1), 24 h (T2), 48 h (T3), and 72 h (T4) post-antibacterial treatment were determined, respectively. Furthermore, the receiver operating characteristic curve (ROC) analysis was performed to evaluate the clinical value of indicators. RESULTS: In IAE group, the IL-6 levels gradually decreased from T0 to T4, and the CRP levels significantly decreased at 48 to 72 h, whereas both IL-6 and CRP remained at high levels in the IAI group. The PCT levels in both groups increased at the early stage of anti-infection (T1-T2) and reached to peak at T1-T2 in effective group. ROC curve analysis identified IL-6 as a predictive biomarker for initial antibiotic efficacy at 12, 48, and 72 h after treatment, with the AUC of 0.698, 0.744, and 0.821, respectively. In addition, CRP demonstrated predictive ability of initial antibiotics against infection at 24, 48, and 72 h after therapy, with the AUC of 0.724, 0.741, and 0.797, respectively. ROC curve analysis of percentage changes demonstrated that IL-6 percentage change showed predictive ability of antibiotic efficacy at the early stage, and both the IL-6 and CRP percentage changes showed the predictive ability of antibiotic efficacy 48 or 72 h after antibiotics therapy. CONCLUSION: This study confirmed IL-6 and CRP levels, and the percentage change in IL-6 as the biomarkers for initial antibiotic efficacy prediction in hematological disorder patients with FN.

Solving the Jigsaw puzzle of phytosterol diversity by a novel sterol methyltransferase from Zea mays.

Phytosterols are vital structural and regulatory components in plants. Zea mays produces a series of phytosterols that are specific to corn. However, the underline biosynthetic mechanism remains elusive. In this study, we identified a novel sterol methyltransferase from Z. mays (ZmSMT1-2) which showed a unique feature compared with documented plant SMTs. ZmSMT1-2 showed a substrate preference for cycloartenol. Using S-adenosyl-L-methionine (AdoMet) as a donor, ZmSMT1-2 converted cycloartenol into alkylated sterols with unique side-chain architectures, including Δ25(27) (i.e., cyclolaudenol and cycloneolitsol) and Δ24(25) (i.e., cyclobranol) sterols. Cycloneolitsol is identified as a product of SMTs for the first time. Our discovery provides a previously untapped mechanism for phytosterol biosynthesis and adds another layer of diversity of sterol biosynthesis.

Serum fat-soluble vitamin levels may be related to sudden sensorineural hearing loss.

BACKGROUND: Few related studies reported yet the association between vitamins and the onset of sudden sensorineural hearing loss (SSNHL). OBJECTIVE: To explore the relationship between the serum levels of fat-soluble vitamins A, D, and E and the risk of SSNHL. METHODS: This retrospective analysis included 310 SSNHL patients and 154 people without risk of hearing loss. The demographic information of all participants like age, gender, body mass index, occupation, cigarette smoking or drinking status, etc. were recorded. The serum levels of vitamins A, D, and E were determined using the electrochemical method. RESULTS: The results indicated that serum vitamin D levels in SSNHL patients were significantly lower. Vitamin D deficiency was only observed in SSNHL group. Similarly, serum vitamin A levels in female SSNHL patients were significantly lower than the control group. Meanwhile, serum vitamin E levels in male SSNHL patients were significantly lower than the control group. CONCLUSION AND SIGNIFICANCE: Our results revealed that the serum levels of fat-soluble vitamins A, D, and E in SSNHL patients were lower than those in the control group with normal hearing, indicating that the decrease of serum fat-soluble vitamins may be related to SSNHL pathogenesis.

Endoscopic type 1 tympanoplasty: comparison of the effects of three different thicknesses grafts.

BACKGROUND: The effects of graft thickness on tympanoplasty is uncertain. OBJECTIVE: To compare the results of endoscopic tympanoplasty using different thicknesses of autologous tissues. METHODS: This retrospective analysis included 186 patients who received type I tympanoplasty, divided into three main groups based of grafting material: perichondrium (A), cartilage-perichondrium (B), or cartilage-perichondrium plus additional perichondrium (C). Group A was subdivided based on whether the placement was inside (A1) or outside (A2) of the malleus. The hearing improvement, graft success rate, and surgery duration were analysed. RESULTS: Statistical analysis showed significant hearing improvement in the three main groups (p < .001); recovery in group A occurred the earliest. Six months postoperatively, group A1 showed significantly greater hearing recovery compared with groups B and C (p < .05). There were no statistical differences the other groups (p > .05) or in the graft success rate among the three main groups (p = .235). The surgery duration of group A was significantly longer than that of groups B and C (p < .001). CONCLUSION AND SIGNIFICANCE: Our results suggest that graft thickness affects hearing recovery; however, graft thickness does not affect the rate of grafting success. Endoscopic transplantation of the perichondrium is more difficult and requires more time.

A Secreted Reporter for Blood Monitoring of Pyroptotic Cell Death.

Pyroptotic cell death is a phenomenon that runs through all life activities and plays an important role in physiological and pathological processes of the body's metabolism. It is of big biological significance to understand the phenomenon and nature of cell pyroptosis. In the process of cell pyroptosis, the pore-forming effector gasdermin D (GSDMD) is cleaved to form oligomers, which are inserted into the cell membrane, causing rapid cell death. However, the effective cell death induced by GSDMD complicates our ability to understand the behavior of pyroptosis. In this work, we performed molecular mutagenesis to develop a genetically encoded pyroptotic reporter, where a secreted Gaussia luciferase (Gluc) was strategically placed in the p30-p20 tolerated junction of GSDMD to support natural pyrophosphorylation and promote live imaging of cell pyroptosis. In addition, we demonstrated that this fused Gluc-GSDMD reporter executed inflammatory body-dependent pyroptosis in response to extracellular stimuli, and that the lysed p30-GSDMD can be secreted out of the cell and can be detected in the culture medium and animal blood. Therefore, our study provides a valuable tool that not only noninvasive and real-time monitoring of cell pyroptosis, but also affords a high-throughput functional screening of pyroptosis-targeted compounds in cultured cells and animal models.

A Functionalized Polydopamine Theranostic Nanoprobe for Efficient Imaging of miRNA-21 and In Vivo Synergetic Cancer Therapy.

MicroRNAs (miRNAs) are emerging as vital biomarkers since their abnormal expression is associated with various disease types including cancer. Therefore, it is essential to develop a sensitive and specific platform to monitor the dynamic expression of miRNAs for early clinical diagnosis and treatment. In this study, we designed a functionalized polydopamine (PDA)-based theranostic nanoprobe for efficient detection of miRNA-21 and in vivo synergistic cancer therapy. PDA was modified with polyethylene glycol (PEG) and the obtained PDA-PEG nanoparticles showed good stability in different solutions. PDA-PEG nanoparticles were loaded with fluorescein isothiocyanate (FITC)-labeled hairpin DNA (hpDNA) and an anticancer drug doxorubicin (DOX). In the absence of miRNA-21, PDA effectively quenched the fluorescence of FITC-labeled hpDNA. The presence of miRNA-21 specifically recognized hpDNA and induced the dissociation of hpDNA from PDA-PEG and subsequently recovered the fluorescence signals. Upon cellular uptake of these nanoprobes, a dose-dependent fluorescence activation and synergetic cytotoxic effect were observed due to the release of DOX and inhibition of miRNA-21 function. Furthermore, PDA-PEG-DOX-hpDNA nanoparticles can afford long-term monitoring of miRNA-21 and combined therapeutic efficacy in the nude mice bearing 4T1 tumors. Our results demonstrate the capability of PDA-PEG-DOX-hpDNA as a theranostic nanoprobe for continuously tracking of miRNAs and synergetic cancer therapy.

p53-dependent upregulation of miR-16-2 by sanguinarine induces cell cycle arrest and apoptosis in hepatocellular carcinoma.

MicroRNAs (miRNAs) were involved in cancer progression, and the targeting of miRNAs by natural agents has opened avenues for cancer treatment and drug development. miR-16 functions as a tumor suppressor and is frequently deleted or downregulated in various human cancers, including hepatocellular carcinoma (HCC). In the present study, we employed a miR-16-responsive luciferase reporter to screen candidate compounds that modulate miR-16 expression from a natural product library. One compound, sanguinarine (SG), was capable of activating miR-16 in HCC cells with wildtype or mutated p53 expression but not in p53-deleted HCC cells. Mechanistic investigations revealed that SG increased p53 occupancy on the miR-16-2 promoter and decreased the expression of miR-16 target genes, including Bcl-2 and cyclin D1. Moreover, SG significantly inhibited HCC cell proliferation in a p53-dependent manner by inducing cell cycle arrest and reactive oxygen species (ROS)-associated apoptosis. Silencing miR-16 by treatment with anti-miR16 miRNA inhibitors rescued the cell viability repression effect caused by SG. Importantly, SG dramatically suppressed tumor growth in an HCC xenograft model, with little cytotoxicity. Taken together, our results provide a preclinical proof-of-concept for SG as a potential strategy for HCC treatment based on the restoration of miR-16 tumor suppressor function.

SOCE mediated by STIM and Orai is essential for pacemaker activity in the interstitial cells of Cajal in the gastrointestinal tract.

Electrical pacemaker activity generates phasic contractions and motility patterns such as segmentation and peristalsis in the gastrointestinal tract. Pacemaker currents are generated in interstitial cells of Cajal (ICC), which release Ca2+ from intracellular stores that stimulates Ca2+-activated Cl- channels (CaCCs) in the plasma membrane. Thus, Ca2+ stores must be maintained to sustain pacemaker activity. Store-operated Ca2+ entry (SOCE) facilitates the refilling of Ca2+ stores by a mechanism dependent upon interactions between STIM and Orai proteins. We investigated the role of SOCE in ICC pacemaker activity. Reintroduction of extracellular Ca2+ in store-depleted ICC resulted in CaCC activation. Blocking CaCCs revealed an inwardly rectifying current with properties of a Ca2+ release-activated current (ICRAC). An inhibitory peptide that interfered with the STIM-Orai interaction blocked ICRAC in HEK 293 cells expressing STIM1 and Orai1 and blocked spontaneous transient inward currents (STICs) and slow wave currents in ICC. STICs, which are fundamental pacemaker events in ICC, were blocked by an Orai antagonist. Imaging of Ca2+ transients linked to pacemaker activity in ICC in intact muscles showed that the Orai antagonist blocked Ca2+ transients in ICC. These data suggest that Ca2+ recovery through STIM-Orai interactions is necessary to maintain ICC pacemaker activity.

The effects of mitochondrial inhibitors on Ca2+ signalling and electrical conductances required for pacemaking in interstitial cells of Cajal in the mouse small intestine.

Interstitial cells of Cajal (ICC-MY) are pacemakers that generate and propagate electrical slow waves in gastrointestinal (GI) muscles. Slow waves appear to be generated by the release of Ca2+ from intracellular stores and activation of Ca2+-activated Cl- channels (Ano1). Conduction of slow waves to smooth muscle cells coordinates rhythmic contractions. Mitochondrial Ca2+ handling is currently thought to be critical for ICC pacemaking. Protonophores, inhibitors of the electron transport chain (FCCP, CCCP or antimycin) or mitochondrial Na+/Ca2+ exchange blockers inhibited slow waves in several GI muscles. Here we utilized Ca2+ imaging of ICC in small intestinal muscles in situ to determine the effects of mitochondrial drugs on Ca2+ transients in ICC. Muscles were obtained from mice expressing a genetically encoded Ca2+ indicator (GCaMP3) in ICC. FCCP, CCCP, antimycin, a uniporter blocker, Ru360, and a mitochondrial Na+/Ca2+ exchange inhibitor, CGP-37157 inhibited Ca2+ transients in ICC-MY. Effects were not due to depletion of ATP, as oligomycin did not affect Ca2+ transients. Patch-clamp experiments were performed to test the effects of the mitochondrial drugs on key pacemaker conductances, Ano1 and T-type Ca2+ (CaV3.2), in HEK293 cells. Antimycin blocked Ano1 and reduced CaV3.2 currents. CCCP blocked CaV3.2 current but did not affect Ano1 current. Ano1 and Cav3.2 currents were inhibited by CGP-37157. Inhibitory effects of mitochondrial drugs on slow waves and Ca2+ signalling in ICC can be explained by direct antagonism of key pacemaker conductances in ICC that generate and propagate slow waves. A direct obligatory role for mitochondria in pacemaker activity is therefore questionable.

Low-dose 5-fluorouracil sensitizes HepG2 cells to TRAIL through TRAIL receptor DR5 and survivin-dependent mechanisms.

Tumour necrosis factor-related apoptosis-inducing ligand (TRAIL) is a promising candidate for cancer treatment due to its highly selective apoptosis-inducing action on tumour cells without harming normal cells. However, because of TRAIL resistance by some cancer cells, combined treatment with sensitizing agents is required to enhance the anticancer potential of TRAIL. In the present study, we investigated the sensitizing effect of 5-fluorouracil (5-FU) on TRAIL-induced apoptosis in TRAIL-resistant HepG2 hepatocarcinoma cells. The results show that 5-FU pretreatment could sensitize HepG2 cells to TRAIL-mediated apoptosis. The enhanced induction of cell death by the 5-FU/TRAIL combination was mediated by DR5 up-regulation and survivin down-regulation. Furthermore, this combination treatment significantly inhibited the growth of human xenografts in vivo. In conclusion, this study demonstrates that the combination of a sensitizing agent and TRAIL may be a novel and effective therapeutic regimen for treating human hepatocellular carcinoma.

N,N'-Dioxide/nickel(ii)-catalyzed asymmetric Diels-Alder reaction of cyclopentadiene with 2,3-dioxopyrrolidines and 2-alkenoyl pyridines.

A chiral N,N'-dioxide/Ni(OTf)2 complex-catalyzed asymmetric Diels-Alder reaction of cyclopentadiene with 2,3-dioxopyrrolidines and 2-alkenoyl pyridines has been achieved. The corresponding chiral bridged compounds were obtained in high yields with excellent dr and ee values (up to 97% yield, 95 : 5 dr and 97% ee).

Nickel(II)-catalyzed asymmetric propargyl and allyl Claisen rearrangements to allenyl- and allyl-substituted ß-ketoesters.

Highly efficient catalytic asymmetric Claisen rearrangements of O-propargyl ß-ketoesters and O-allyl ß-ketoesters have been accomplished under mild reaction conditions. In the presence of the chiral N,N'-dioxide/Ni(II) complex, a wide range of allenyl/allyl-substituted all-carbon quaternary ß-ketoesters was obtained in generally good yield (up to 99%) and high diastereoselectivity (up to 99:1 d.r.) with excellent enantioselectivity (up to 99% ee).

Laminar shear stress upregulates endothelial Ca²âº-activated K⁺ channels KCa2.3 and KCa3.1 via a Ca²âº/calmodulin-dependent protein kinase kinase/Akt/p300 cascade.

In endothelial cells (ECs), Ca²âº-activated K⁺ channels KCa2.3 and KCa3.1 play a crucial role in the regulation of arterial tone via producing NO and endothelium-derived hyperpolarizing factors. Since a rise in intracellular Ca²âº levels and activation of p300 histone acetyltransferase are early EC responses to laminar shear stress (LS) for the transcriptional activation of genes, we examined the role of Ca²âº/calmodulin-dependent kinase kinase (CaMKK), the most upstream element of a Ca²âº/calmodulin-kinase cascade, and p300 in LS-dependent regulation of KCa2.3 and KCa3.1 in ECs. Exposure to LS (15 dyn/cm²) for 24 h markedly increased KCa2.3 and KCa3.1 mRNA expression in cultured human coronary artery ECs (3.2 ± 0.4 and 45 ± 10 fold increase, respectively; P < 0.05 vs. static condition; n = 8-30), whereas oscillatory shear (OS; ± 5 dyn/cm² × 1 Hz) moderately increased KCa3.1 but did not affect KCa2.3. Expression of KCa2.1 and KCa2.2 was suppressed under both LS and OS conditions, whereas KCa1.1 was slightly elevated in LS and unchanged in OS. Inhibition of CaMKK attenuated LS-induced increases in the expression and channel activity of KCa2.3 and KCa3.1, and in phosphorylation of Akt (Ser473) and p300 (Ser1834). Inhibition of Akt abolished the upregulation of these channels by diminishing p300 phosphorylation. Consistently, disruption of the interaction of p300 with transcription factors eliminated the induction of these channels. Thus a CaMKK/Akt/p300 cascade plays an important role in LS-dependent induction of KCa2.3 and KCa3.1 expression, thereby regulating EC function and adaptation to hemodynamic changes.

A functional role for the 'fibroblast-like cells' in gastrointestinal smooth muscles.

Smooth muscles, as in the gastrointestinal tract, are composed of several types of cells. Gastrointestinal muscles contain smooth muscle cells, enteric neurons, glial cells, immune cells, and various classes of interstitial cells. One type of interstitial cell, referred to as 'fibroblast-like cells' by morphologists, are common, but their function is unknown. These cells are found near the terminals of enteric motor neurons, suggesting they could have a role in generating neural responses that help control gastrointestinal movements. We used a novel mouse with bright green fluorescent protein expressed specifically in the fibroblast-like cells to help us identify these cells in the mixture of cells obtained when whole muscles are dispersed with enzymes. We isolated these cells and found they respond to a major class of inhibitory neurotransmitters - purines. We characterized these responses, and our results provide a new hypothesis about the role of fibroblast-like cells in smooth muscle tissues.

TNF-alpha inhibits the CD3-mediated upregulation of voltage-gated K+ channel (Kv1.3) in human T cells.

A long term treatment of T cells with tumor necrosis factor alpha (TNF-alpha) paradoxically inhibits the immunologic responses to TCR/CD3 stimulation. The voltage-gated K(+) channels (K(v)) of T cells attracted attention as a pharmacological target for the treatment of autoimmune diseases. Here, the authors investigated the effects of TNF-alpha on the K(v) current (I(Kv)) and its upregulation by CD3 in human T cells. Acute treatment with TNF-alpha (10 min) temporarily decreased I(Kv) in Jurkat-T cells (cells subsequently recovered after treatment >12h), whereas CD3 stimulation for 24h increased I(Kv) amplitude more than two-fold. Furthermore, chronic pretreatment with TNF-alpha almost completely blocked the I(Kv) increase induced by CD3 stimulation. An immunoblot study confirmed an increase in the protein level of K(v) induced by CD3 stimulation, and its inhibition by TNF-alpha pretreatment. In addition, the facilitation of I(Kv) by CD3 stimulation and its inhibition by pretreatment with TNF-alpha were confirmed in freshly isolated human peripheral CD4(+) T cells, in which the voltage-dependence of I(Kv) was unaffected by TNF-alpha and/or CD3 stimulation. We conclude that the inhibition of CD3-induced K(v) upregulation by TNF-alpha might be associated with the paradoxical suppression of T cell function by TNF-alpha under conditions of chronic inflammation.

Identification of the large-conductance background K+ channel in mouse B cells as TREK-2.

Mouse B cells and their cell line (WEHI-231) express large-conductance background K(+) channels (LK(bg)) that are activated by arachidonic acids, characteristics similar to TREK-2. However, there is no evidence to identify the molecular nature of LK(bg); some properties of LK(bg) were partly different from the reported results of TREK type channels. In this study, we compared the properties of cloned TREK-2 and LK(bg) in terms of their sensitivities to ATP, phosphatidylinositol 4,5-bisphosphate (PIP(2)), intracellular pH (pH(i)), and membrane stretch. Similar to the previous findings of LK(bg), TREK-2 showed spontaneous activation after membrane excision (i-o patch) and were inhibited by MgATP or by PIP(2). The inhibition by MgATP was prevented by wortmannin, suggesting membrane-delimited regulation of TREKs by phosphoinositide (PI) kinase. The same was observed with the property of LK(bg); the activation of TREK-2 by membrane stretch was suppressed by U73122 (PLC inhibitor). As with the known properties of TREK-2, LK(bg) were activated by acidic pH(i) and inhibited by PKC activator. Finally, we confirmed the expression of TREK-2 in WEHI-231 by using RT-PCR and immunoblot analyses. The amplitude of background K(+) current and the TREK-2 expression in WEHI-231 were commonly decreased by genetic knockdown of TREK-2 using small interfering RNA. The downregulation of TREK-2 attenuated Ca(2+)-influx induced by arachidonic acid in WEHI-231. As a whole, these results strongly indicate that TREK-2 encodes LK(bg) in mouse B cells. We also newly suggest that the low activity of TREK-2 in intact cells is due to the inhibition by intrinsic PIP(2).

Role of aryl hydrocarbon receptor nuclear translocator in KATP channel-mediated insulin secretion in INS-1 insulinoma cells.

Aryl hydrocarbon receptor nuclear translocator (ARNT) has been known to participate in cellular responses to xenobiotic and hypoxic stresses, as a common partner of aryl hydrocarbon receptor and hypoxia inducible factor-1/2alpha. Recently, it was reported that ARNT is essential for adequate insulin secretion in response to glucose input and that its expression is downregulated in the pancreatic islets of diabetic patients. In the present study, the authors addressed the mechanism by which ARNT regulates insulin secretion in the INS-1 insulinoma cell line. In ARNT knock-down cells, basal insulin release was elevated, but insulin secretion was not further stimulated by a high-glucose challenge. Electrophysiological analyses revealed that glucose-dependent membrane depolarization was impaired in these cells. Furthermore, K(ATP) channel activity and expression were reduced. Of two K(ATP) channel subunits, Kir6.2 was found to be positively regulated by ARNT at the mRNA and protein levels. Based on these results, the authors suggest that ARNT expresses K(ATP) channel and by so doing regulates glucose-dependent insulin secretion.