A deep-learning model using enhanced chest CT images to predict PD-L1 expression in non-small-cell lung cancer patients.

AIM: To develop a deep-learning model using contrast-enhanced chest computed tomography (CT) images to predict programmed death-ligand 1 (PD-L1) expression in patients with non-small-cell lung cancer (NSCLC). MATERIALS AND METHODS: Preoperative enhanced chest CT images and immunohistochemistry results for PD-L1 expression (<1% and ≥1% were defined as negative and positive, respectively) were collected retrospectively from 125 NSCLC patients to train and validate a deep-learning radiomics model (DLRM) for the prediction of PD-L1 expression in tumours. The DLRM was developed by combining the deep-learning signature (DLS) obtained from a convolutional neural network and clinicopathological factors. The indexes of the area under the curve (AUC), integrated discrimination improvement (IDI), and decision curve analysis (DCA) were used to evaluate the efficiency of the DLRM. RESULTS: DLS and tumour stage were identified as independent predictors of PD-L1 expression by the DLRM. The AUCs of the DLRM were 0.804 (95% confidence interval: 0.697-0.911) and 0.804 (95% confidence interval: 0.679-0.929) in the training and validation cohorts, respectively. IDI analysis showed the DLRM had better diagnostic accuracy than DLS (0.0028 [p<0.05]) in the validation cohort. Additionally, DCA revealed that the DLRM had more net benefit than the DLS for clinical utility. CONCLUSION: The proposed DLRM using enhanced chest CT images could function as a non-invasive diagnostic tool to differentiate PD-L1 expression in NSCLC patients.

Apelin-13 promotes late endothelial progenitor cells differentiation by regulating Krüppel-like factor 4.

OBJECTIVE: The endothelial progenitor cells (EPCs) differentiation plays an essential role in angiogenesis. The purpose of this study is to determine the potential mechanisms of apelin-13 in EPCs differentiation. MATERIALS AND METHODS: The rats EPCs were isolated from the bone marrow and identified by DIL-acLDL and FITC-UEA-1 staining. EPCs were divided into four groups: the negative control group, the Krüppel-like factor 4 (KLF4) upregulation group, the KLF4 downregulation group, and the apelin-13 group. The EPCs differentiation was evaluated by cell migration, proliferation, and the expressions of surface markers CD31, CD34, CD133, and VEGFR-2 on mRNA and the protein levels, as well as immunofluorescence. RESULTS: In the KLF4 up-regulation and apelin-13 groups, the EPCs number of G1, S, and G2/M phases decreased and suggested that KLF4 and apelin-13 were closely related to the EPCs proliferation. EPCs showed stronger migration ability with the elevation of KLF4 and apelin-13 while declined migration was detected in KLF4 siRNA transfected EPCs. The surface markers expressions on mRNA and the protein levels increased in the KLF4 upregulation group, and in the apelin-13 group there were similar results, as well as increased KLF4 expression. CONCLUSIONS: The upregulation of apelin-13 significantly increased the expressions of EPCs surface markers, which were involved in early EPCs differentiated into late EPCs and influenced the cells migration and proliferation. Taking the elevation of KLF4 which performed similar effects of apelin-13, we believe that apelin-13 activates or synergizes with KLF4 to promote this process.

Etomidate uniquely modulates the desensitization of recombinant α1ß3δ GABA(A) receptors.

Central GABA(A) receptors mediate GABAergic phasic and tonic inhibition. While synaptic αßγ GABA(A) receptors primarily mediate phasic inhibition, extrasynaptic αßδ receptors play an important role in mediating tonic inhibition. Etomidate is a general anesthetic that produces its effects by enhancing GABA(A) receptor activity. We previously showed that etomidate modulates the gating of oocyte-expressed αßγ and αßδ receptors with similar overall allosteric impact, but different pharmacological patterns. In αßγ receptors, etomidate enhances apparent GABA sensitivity (reduces GABA EC50), modestly increases maximal GABA efficacy, and slows current deactivation without affecting desensitization (Zhong et al., 2008). In αßδ receptors characterized by low GABA efficacy, etomidate dramatically increases responses to both low and maximal GABA. The effects of etomidate on desensitization and deactivation of αßδ receptors are unknown. To investigate the kinetic effects of etomidate on α1ß3δ receptors of defined subunit arrangement, we expressed concatenated trimer (ß3-α1-δ) and dimer (ß3-α1) GABA(A) receptor subunit assemblies in human embryonic kidney (HEK)293T cells and recorded whole-cell voltage-clamp currents during rapid external solution exchanges. As expected, etomidate substantially increased maximal GABA-induced currents and prolonged deactivation. Moreover, desensitization was significantly decreased by etomidate. During prolonged GABA applications, etomidate enhanced steady-state currents more than peak currents. Thus, etomidate enhances tonic GABAergic inhibition through extrasynaptic αßδ receptors by both augmenting gating and reducing desensitization.

Association of peroxisome proliferator-activated receptor-delta polymorphisms with sugar metabolism indices and tumor necrosis factor alpha level.

This study aims to investigate the association of peroxisome proliferator-activated receptor (PPAR) delta -87T/C polymorphism with several sugar metabolism indices and tumor necrosis factor α (TNF α) level. The body mass index (BMI), waist size, and levels of fasting plasma glucose, serum lipid, fasting insulin, TNFα, and PPAR delta -87T/C of 286 patients with type 2 diabetes mellitus (T2DM) and 158 subjects with normal fasting glucose (NFG) were measured in a Dalian population. The distribution of genotypic frequencies between T2DM and NFG were not significantly different (χ(2) = 0.012, P = 0.994). BMI, fasting blood glucose (FBG), homeostasis model assessment-estimated insulin resistance (HOMA-IR), triglyceride, and TNFα levels were significantly different among different T2DM genotypes. HOMA-IR and FBG were significantly different among different NFG genotypes. The PPAR delta -87T/C polymorphism is known to be closely related with glucose levels and lipid metabolism. A close relationship was also found between HOMA-IR and TNFα levels and HOMA-IR and FBG in T2DM and NFG, respectively.

Global microRNA profiles and signaling pathways in the development of cardiac hypertrophy

Hypertrophy is a major predictor of progressive heart disease and has an adverse prognosis. MicroRNAs (miRNAs) that accumulate during the course of cardiac hypertrophy may participate in the process. However, the nature of any interaction between a hypertrophy-specific signaling pathway and aberrant expression of miRNAs remains unclear. In this study, Spague Dawley male rats were treated with transverse aortic constriction (TAC) surgery to mimic pathological hypertrophy. Hearts were isolated from TAC and sham operated rats (n=5 for each group at 5, 10, 15, and 20 days after surgery) for miRNA microarray assay. The miRNAs dysexpressed during hypertrophy were further analyzed using a combination of bioinformatics algorithms in order to predict possible targets. Increased expression of the target genes identified in diverse signaling pathways was also analyzed. Two sets of miRNAs were identified, showing different expression patterns during hypertrophy. Bioinformatics analysis suggested the miRNAs may regulate multiple hypertrophy-specific signaling pathways by targeting the member genes and the interaction of miRNA and mRNA might form a network that leads to cardiac hypertrophy. In addition, the multifold changes in several miRNAs suggested that upregulation of rno-miR-331*, rno-miR-3596b, rno-miR-3557-5p and downregulation of rno-miR-10a, miR-221, miR-190, miR-451 could be seen as biomarkers of prognosis in clinical therapy of heart failure. This study described, for the first time, a potential mechanism of cardiac hypertrophy involving multiple signaling pathways that control up- and downregulation of miRNAs. It represents a first step in the systematic discovery of miRNA function in cardiovascular hypertrophy.

Global microRNA profiles and signaling pathways in the development of cardiac hypertrophy.

Hypertrophy is a major predictor of progressive heart disease and has an adverse prognosis. MicroRNAs (miRNAs) that accumulate during the course of cardiac hypertrophy may participate in the process. However, the nature of any interaction between a hypertrophy-specific signaling pathway and aberrant expression of miRNAs remains unclear. In this study, Spague Dawley male rats were treated with transverse aortic constriction (TAC) surgery to mimic pathological hypertrophy. Hearts were isolated from TAC and sham operated rats (n=5 for each group at 5, 10, 15, and 20 days after surgery) for miRNA microarray assay. The miRNAs dysexpressed during hypertrophy were further analyzed using a combination of bioinformatics algorithms in order to predict possible targets. Increased expression of the target genes identified in diverse signaling pathways was also analyzed. Two sets of miRNAs were identified, showing different expression patterns during hypertrophy. Bioinformatics analysis suggested the miRNAs may regulate multiple hypertrophy-specific signaling pathways by targeting the member genes and the interaction of miRNA and mRNA might form a network that leads to cardiac hypertrophy. In addition, the multifold changes in several miRNAs suggested that upregulation of rno-miR-331*, rno-miR-3596b, rno-miR-3557-5p and downregulation of rno-miR-10a, miR-221, miR-190, miR-451 could be seen as biomarkers of prognosis in clinical therapy of heart failure. This study described, for the first time, a potential mechanism of cardiac hypertrophy involving multiple signaling pathways that control up- and downregulation of miRNAs. It represents a first step in the systematic discovery of miRNA function in cardiovascular hypertrophy.

Etomidate produces similar allosteric modulation in α1ß3δ and α1ß3γ2L GABA(A) receptors.

BACKGROUND AND PURPOSE: Neuronal GABA(A) receptors are pentameric chloride ion channels, which include synaptic αßγ and extrasynaptic αßδ isoforms, mediating phasic and tonic inhibition respectively. Although the subunit arrangement of αßγ receptors is established as ß-α-γ-ß-α, that of αßδ receptors is uncertain and possibly variable. We compared receptors formed from free α1, ß3 and δ or γ2L subunits and concatenated ß3-α1-δ and ß3-α1 subunit assemblies (placing δ in the established γ position) by investigating the effects of R-(+)-etomidate (ETO), an allosteric modulator that selectively binds to transmembrane interfacial sites between ß3 and α1. EXPERIMENTAL APPROACH: GABA-activated receptor-mediated currents in Xenopus oocytes were measured electrophysiologically, and ETO-induced allosteric shifts were quantified using an established model. KEY RESULTS: ETO (3.2 µM) similarly enhanced maximal GABA (1 mM)-evoked currents in oocytes injected with 5 ng total mRNA and varying subunit ratios, for α1ß3(1:1), α1ß3δ(1:1:1) and α1ß3δ(1:1:3), but this potentiation by ETO was significantly greater for ß3-α1-δ/ß3-α1(1:1) receptors. Reducing the amount of α1ß3δ(1:1:3) mRNA mixture injected (0.5 ng) increased the modulatory effect of ETO, matching that seen with ß3-α1-δ/ß3-α1(1:1, 1 ng). ETO similarly reduced EC50s and enhanced maxima of GABA concentration-response curves for both α1ß3δ and ß3-α1-δ/ß3-α1 receptors. Allosteric shift parameters derived from these data depended on estimates of maximal GABA efficacy, and the calculated ranges overlap with allosteric shift values for α1ß3γ2L receptors. CONCLUSION AND IMPLICATIONS: Reducing total mRNA unexpectedly increased δ subunit incorporation into receptors on oocyte plasma membranes. Our results favour homologous locations for δ and γ2L subunits in α1ß3γ2/δ GABA(A) receptors.

Achieving synaptically relevant pulses of neurotransmitter using PDMS microfluidics.

Fast synaptic transmission is mediated by post-synaptic ligand-gated ion channels (LGICs) transiently activated by neurotransmitter released from pre-synaptic vesicles. Although disruption of synaptic transmission has been implicated in numerous neurological and psychiatric disorders, effective and practical methods for studying LGICs in vitro under synaptically relevant conditions are unavailable. Here, we describe a novel microfluidic approach to solution switching that allows for precise temporal control over the neurotransmitter transient while substantially increasing experimental throughput, flexibility, reproducibility, and cost-effectiveness. When this system was used to apply ultra-brief ( approximately 400micros) GABA pulses to recombinant GABA(A) receptors, members of the cys-loop family of LGICs, the resulting currents resembled hippocampal inhibitory post-synaptic currents (IPSCs) and differed from currents evoked by longer, conventional pulses, illustrating the importance of evaluating LGICs on a synaptic timescale. This methodology should therefore allow the effects of disease-causing mutations and allosteric modulators to be evaluated in vitro under physiologically relevant conditions.

Expression and pathological relevance of inducible nitric oxide synthase in osteosarcoma of the jaws.

The aim of the present study was to examine the expression of inducible nitric oxide synthase (iNOS) in osteosarcoma of the jaw, and its relationship with tumour angiogenesis and clinicopathological characteristics. Streptavidin peroxidase immunohistochemical staining was used to detect the expression level of iNOS and CD34 in paraffin-embedded samples from 25 patients. Osteosarcoma of the jaw was associated with overexpression of iNOS, which correlated with tumour microvessel density (MVD). iNOS expression correlated with the size, pathological grade and clinical stage of the osteosarcoma, and also with clinicopathological characteristics such as primary occurrence or recurrence of tumours. There was no correlation with metastasis. iNOS may promote tumour angiogenesis in osteosarcoma of the jaw, and so may represent an important target in anti-tumour therapy.

Glutamatergic activation of the amygdala differentially mimics the effects of audiogenic seizure kindling in two substrains of genetically epilepsy-prone rats.

Comparisons of neuronal network mechanisms in closely related inherited seizure models are providing novel insights into epileptogenic pathophysiology. Genetically epilepsy-prone rats (GEPRs) exist in two substrains that inherit long-term susceptibility to behaviorally distinct audiogenic seizures (AGS). GEPR-3s exhibit generalized clonic AGS, while GEPR-9s exhibit generalized tonic AGS. After AGS kindling the tonic AGS of GEPR-9s is followed by generalized posttonic clonus (PTC), while the generalized clonic AGS is followed by facial and forelimb (F&F) clonus in GEPR-3s. PTC and F&F clonus are very rare in GEPRs before AGS kindling. The neuronal network subserving AGS in GEPR-9s lies exclusively in brainstem sites, but amygdala (AMG) and other sites are recruited into the network after AGS kindling. The present study attempted to mimic the effects of AGS kindling by bilaterally microinjecting subconvulsive doses of N-methyl-D-aspartate (NMDA) into the AMG of nonkindled GEPRs. NMDA (10 nmol/side) microinjected into AMG reversibly induced susceptibility to F&F clonus immediately following generalized clonic AGS in most nonkindled GEPR-3s. NMDA (7.5 nmol/side), microinjected into AMG temporarily induced susceptibility to generalized PTC immediately following tonic AGS in most nonkindled GEPR-9s. No seizures were induced in normal rats by these treatments, and no seizures were seen in GEPRs with these NMDA doses except those induced by acoustic stimuli. These findings support a critical role in AGS kindling for the AMG in the neuronal networks for both forms of AGS. However, the behavioral effect of the treatment was different in the two AGS substrains, suggesting interrelated but not identical pathophysiological mechanisms in these closely related epilepsy models.

Modulation of audiogenically kindled seizures by gamma-aminobutyric acid-related mechanisms in the amygdala.

Repetitive induction of audiogenic seizures (AGSs) ("AGS kindling") results in expansion of the AGS neuronal network from the brainstem to forebrain structures. AGSs in kindled genetically epilepsy-prone rats (GEPR-9s) exhibit a significant increase in the duration of posttonic clonus (PTC). The amygdala (AMG) does not appear to be a required network component before AGS kindling, but this structure is implicated in the seizure network after AGS kindling. gamma-Aminobutyric acid (GABA) is a major neurotransmitter in AMG, and histamine receptor activation is also reported to stimulate GABA release. The present study examined the effect on audiogenically kindled seizures of focal microinjections into the AMG of GEPR-9s. AGS kindling involved induction of 14 AGSs in GEPR-9s. Bilateral microinjection of a GABA(A) agonist, muscimol (0.3 nmol/side), into the AMG significantly reduced the duration of PTC, starting 0.5 h after drug infusion, with recovery by 24 h. Microinjection of histamine (60 nmol/side) suppressed PTC at 0.5 h, with total blockade at 24 h, but the seizure pattern did not revert to that observed before kindling until 120 h. This long duration suggests that mechanisms in addition to modulation of GABA function may be involved in the effect of histamine. The wild running and tonic components of AGS were never affected by microinjection of these agents into the AMG. These findings confirm previous work suggesting that the AMG is not a required nucleus in the AGS neuronal network before kindling. However, the AMG becomes critical in expansion of the seizure network during AGS kindling, and audiogenically kindled seizures are negatively modulated by increased GABA function.

Modulation of audiogenic seizures by histamine and adenosine receptors in the inferior colliculus.

Susceptibility to behaviorally similar audiogenic seizures (AGS) occurs genetically and is inducible during ethanol withdrawal (ETX). Comparisons between AGS mechanisms of genetically epilepsy-prone rats (GEPR-9s) and ethanol-withdrawn rats (ETX-Rs) are yielding information about general pathophysiological mechanisms of epileptogenesis. The inferior colliculus (IC) is the AGS initiation site. Excitatory amino acid (EAA) abnormalities in the IC are implicated in AGS, and histamine and adenosine receptor activation each reduce EAA release and inhibit several seizure types. Previous studies indicate that focal infusion of an adenosine receptor agonist into the IC blocked AGS in GEPR-9s, but the effects of adenosine receptor activation in the IC on AGS in ETX-Rs are unknown. The effects of histamine receptor activation on either form of AGS are also unexamined. The present study evaluated effects of histamine or a nonselective adenosine A(1) agonist, 2-chloroadenosine, on AGS by focal microinjection into the IC. Ethanol dependence and AGS susceptibility were induced in normal rats by intragastric ethanol. Histamine (40 or 60 nmol/side) significantly reduced AGS in GEPR-9s, but histamine in doses up to 120 nmol/side did not affect AGS in ETX-Rs. 2-Chloroadenosine (5 or 10 nmol/side) did not affect AGS in ETX-Rs, despite the effectiveness of lower doses of this agent in GEPR-9s reported previously. Thus, histamine and adenosine receptors in the IC modulate AGS of GEPR-9s, but do not modulate ETX-induced AGS. The reasons for this difference may involve the chronicity of AGS susceptibility in GEPR-9s, which may lead to more extensive neuromodulation as compensatory mechanisms to limit the seizures compared to the acute AGS of ETX-Rs.

The induction effect of rifampicin on activity of mephenytoin 4'-hydroxylase related to M1 mutation of CYP2C19 and gene dose.

AIMS: To determine the induction effect of rifampicin on the activity of 4'-hydroxylase in poor metabolizers (PMs) with m1 mutation of S-mephenytoin 4'-hydroxylation and the relationship of the effect with gene dose. METHODS: Seven extensive metabolizers (EMs) of S-mephenytoin 4'-hydroxylation and five PMs with m1 mutation were chosen to take rifampicin 300 mg day(-1) orally for 22 days. Prior to and after rifampicin treatment, each subject was given racemic mephenytoin 100 mg. The 4'-hydroxymephenytoin (4'-OH-MP) excreted in the 0-24 h urine and mephenytoin S/R ratio in the 0-8 h urine were determined by h.p.l.c. and GC, respectively. RESULTS: In all EMs, the excretion of 4'-OH-MP in the 0-24 h urine was increased by 146.4 +/- 17.9%, 0-8 h urinary mephenytoin S/R ratio was decreased by 77.3 +/- 8.8%, the percentage increase in the 0-24 h excretion of 4'-OH-MP in those CYP2C19 homozygous (wt/wt) was greater than that in those heterozygous (wt/m1 and wt/m2) (203.9 +/- 42.5% vs 69.6 +/- 4.1%). 0-8 h urinary mephenytoin S/R ratio of those PMs with m1 mutation was decreased by 9.6%, the amount of 4'-OH-MP excreted in the 0-24 h urine was increased by 80.1 +/- 48.0%. CONCLUSIONS: The activity of 4'-hydroxylase of PMs with m1 mutation of S-mephenytoin 4'-hydroxylation can be induced by rifampicin and the inducing effect of rifampicin on 4'-hydroxylase is gene dependent.