A competing risk predictive model for kidney failure in patients with advanced chronic kidney disease.

BACKGROUND/PURPOSE: Predictive modeling aids in identifying patients at high risk of adverse events. Using routinely collected data, we report a competing risk prediction model for kidney failure. METHODS: A total of 5138 patients with CKD stages 3b-5 were included and randomized into the development and validation cohorts at a ratio of 7:3. The outcome was end-stage kidney disease, defined as the initiation of dialysis or kidney transplantation. All patients were followed-up until December 31, 2020. A Fine and Gray model was applied to estimate the sub-hazard ratio of kidney failure, with death as a competing event. RESULTS: In the development cohort, the mean age was 67.6 ± 13.9 years and 60 % were male. The mean index eGFR and median urinary protein-creatinine ratio (UPCR) were 26.5 ± 12.8 mL/min/1.73 m2 and 1051 mg/g, respectively. The median follow-up duration was 1051 days. The proportion of patients with kidney failure and death was 25.4 % and 14.1 %, respectively. Four models were applied, including eGFR, age, sex, UPCR, systolic and diastolic blood pressure, serum albumin, phosphate, uric acid, haemoglobin, and potassium levels had the best goodness of fit. All models had good discrimination with time-to-event c statistics of 0.89-0.95 in the development cohort and 0.86-0.95 in the validation cohort. The prediction models showed excellent and fairly good calibration at 2 and 5-year risk, respectively. CONCLUSION: Using real-world data, our competing risk model can accurately predict progression to kidney failure over 2 years in patients with advanced CKD.

CART Peptides Differently Regulate Firing Rates and GABAergic Synaptic Inputs of DMV Neurons Innervating the Stomach Antrum and Cecum of Adult Male Rats.

BACKGROUND/AIM: Central administration of cocaine- and amphetamine-regulated transcript peptides (CARTp) alters gastrointestinal motility and reduces food intake in rats. Since neurons in the dorsal motor nucleus of the vagus (DMV) receive GABAergic and glutamatergic inputs and innervate the smooth muscle of gastrointestinal organs, we hypothesized that CARTp acts on the DMV or presynaptic neurons. METHODS: We used 3,3'-dioctadecyloxa-carbocyanine perchlorate (DiO) retrograde tracing with electrophysiological methods to record DMV neurons innervating the stomach antrum or cecum in brainstem slices from adult rats. RESULTS: DiO application did not change the electrophysiological properties of DMV neurons. CART55-102 had no effect on the basal firing rates of neurons in either the stomach antrum-labeled group (SLG) or cecum-labeled group (CLG). When presynaptic inputs were blocked, CART55-102 further increased the firing rates of the SLG, suggesting a direct excitatory effect. Spontaneous inhibitory postsynaptic currents (sIPSCs) occurred at a higher frequency in SLG neurons than in CLG neurons. CART55-102 reduced the amplitude and the frequency of sIPSCs in SLG neurons dose-dependently, with higher doses also reducing spontaneous excitatory postsynaptic currents (sEPSCs). Higher doses of CART55-102 reduced sIPSC and sEPSC amplitudes in CLG neurons, suggesting a postsynaptic effect. In response to incremental current injections, the SLG neurons exhibited less increases in firing activity. Simultaneous applications of current injections and CART55-102 decreased the firing activity of the CLG. Therefore, stomach antrum-projecting DMV neurons possess a higher gating ability to stabilize firing activity. CONCLUSION: The mechanism by which CARTp mediates anorectic actions may be through a direct reduction in cecum-projecting DMV neuron excitability and, to a lesser extent, that of antrum-projecting DMV neurons, by acting on receptors of these neurons.

Differential modulation of avian ß-defensin and Toll-like receptor expression in chickens infected with infectious bronchitis virus.

The host innate immune response either clears invading viruses or allows the adaptive immune system to establish an effective antiviral response. In this study, both pathogenic (passage 3, P3) and attenuated (P110) infectious bronchitis virus (IBV) strains were used to study the immune responses of chicken to IBV infection. Expression of avian ß-defensins (AvBDs) and Toll-like receptors (TLRs) in 16 tissues of chicken were compared at 7 days PI. The results showed that P3 infection upregulated the expression of AvBDs, including AvBD2, 4, 5, 6, 9, and 12, while P110 infection downregulated the expression of AvBDs, including AvBD3, 4, 5, 6, and 9 in most tissues. Meanwhile, the expression level of several TLRs showed a general trend of upregulation in the tissues of P3-infected chickens, while they were downregulated in the tissues of P110-infected chickens. The result suggested that compared with the P110 strain, the P3 strain induced a more pronounced host innate immune response. Furthermore, we observed that recombinant AvBDs (including 2, 6, and 12) demonstrated obvious anti-viral activity against IBV in vitro. Our findings contribute to the proposal that IBV infection induces an increase in the messenger RNA (mRNA) expression of some AvBDs and TLRs, which suggests that AvBDs may play significant roles in the resistance of chickens to IBV replication.

Developmental increase in hippocampal endocannabinoid mobilization: role of metabotropic glutamate receptor subtype 5 and phospholipase C.

Endocannabinoids (eCBs) released from postsynaptic neurons mediate retrograde suppression of neurotransmitter release at central synapses. eCBs are crucial for establishing proper synaptic connectivity in the developing nervous system. Mobilization of eCBs is driven either by a rise in intracellular Ca(2+) (depolarization-induced suppression of inhibition, DSI) or postsynaptic G protein-coupled receptors (GPCRs) that activate phospholipase C beta (PLCß). To determine whether eCB mobilization changes between neonatal and juvenile ages, we used whole cell voltage-clamp recordings of CA1 neurons from rat hippocampal slices at postnatal days 1-18 (neonatal) and 19-43 (juvenile), because many neurophysiological parameters change dramatically between approximately postnatal days 18-20. We found that DSI was slightly greater in juveniles than in neonates, while eCB mobilization stimulated by GPCRs was unchanged. However, when DSI was elicited during GPCR activation, its increase was much greater in juveniles, suggesting that eCB mobilization caused by the synergy between the Ca(2+) and GPCR pathways is developmentally upregulated. Western blotting revealed significant increases in both metabotropic type glutamate receptor 5 (mGluR5) and PLCß1 proteins in juveniles compared with neonates. Responses to pharmacological activation or inhibition of PLC implied that eCB upregulation is associated with a functional increase in PLC activity. We conclude that synergistic eCB mobilization in hippocampal CA1 neurons is greater in juveniles than in neonates, and that this may result from increases in the mGluR5-PLCß1 eCB pathway. The data enhance our understanding of the developmental regulation of the eCB system and may provide insight into diseases caused by improper cortical wiring, or the impact of cannabis exposure during development.

Involvement of dopamine D2 receptor in the diurnal changes of tuberoinfundibular dopaminergic neuron activity and prolactin secretion in female rats.

BACKGROUND: An endogenous dopaminergic (DA) tone acting on D3 receptors has been shown to inhibit tuberoinfundibular (TI) DA neuron activity and stimulate prolactin (PRL) surge in the afternoon of estrogen-primed ovariectomized (OVX+E2) rats. Whether D2 receptor (D2R) is also involved in the regulation of TIDA and PRL rhythms was determined in this study. RESULTS: Intracerebroventricular (icv) injection of PHNO, a D2R agonist, in the morning inhibited TIDA and midbrain DA neurons' activities, and stimulated PRL secretion. The effects of PHNO were significantly reversed by co-administration of raclopride, a D2R antagonist. A single injection of raclopride at 1200 h significantly reversed the lowered TIDA neuron activity and the increased serum PRL level at 1500 h. Dopamine D2R mRNA expression in medial basal hypothalamus (MBH) exhibited a diurnal rhythm, i.e., low in the morning and high in the afternoon, which was opposite to that of TIDA neuron activity. The D2R rhythm was abolished in OVX+E2 rats kept under constant lighting but not in OVX rats with regular lighting exposures. Pretreatment with an antisense oligodeoxynucleotides (AODN, 10 µg/3 µl/day, icv) against D2R mRNA for 2 days significantly reduced D2R mRNAs in central DA neurons, and reversed both lowered TIDA neuron activity and increased serum PRL level in the afternoon on day 3. A diurnal rhythm of D2R mRNA expression was also observed in midbrain DA neurons and the rhythm was significantly knocked down by the AODN pretreatment. CONCLUSIONS: We conclude that a diurnal change of D2R mRNA expression in MBH may underlie the diurnal rhythms of TIDA neuron activity and PRL secretion in OVX+E2 rats.

Unveiling the spatiotemporal dynamics of membrane fouling: A focused review on dynamic fouling characterization techniques and future perspectives.

Membrane technology has emerged as a crucial method for obtaining clean water from unconventional sources in the face of water scarcity. It finds wide applications in wastewater treatment, advanced treatment, and desalination of seawater and brackish water. However, membrane fouling poses a huge challenge that limits the development of membrane-based water treatment technologies. Characterizing the dynamics of membrane fouling is crucial for understanding its development, mechanisms, and effective mitigation. Instrumental techniques that enable in situ or real-time characterization of the dynamics of membrane fouling provide insights into the temporal and spatial evolution of fouling, which play a crucial role in understanding the fouling mechanism and the formulation of membrane control strategies. This review consolidates existing knowledge about the principal advanced instrumental analysis technologies employed to characterize the dynamics of membrane fouling, in terms of membrane structure, morphology, and intermolecular forces. Working principles, applications, and limitations of each technique are discussed, enabling researchers to select appropriate methods for their specific studies. Furthermore, prospects for the future development of dynamic characterization techniques for membrane fouling are discussed, underscoring the need for continued research and innovation in this field to overcome the challenges posed by membrane fouling.

Investigation of the miRNA-mRNA Regulatory Circuits and Immune Signatures Associated with Bronchopulmonary Dysplasia.

Background: Bronchopulmonary dysplasia (BPD) has become a major cause of morbidity and mortality in preterm infants worldwide, yet its pathogenesis and underlying mechanisms remain poorly understood. The present study sought to explore microRNA-mRNA regulatory networks and immune cells involvement in BPD through a combination of bioinformatic analysis and experimental validation. Methods: MicroRNA and mRNA microarray datasets were obtained from the Gene Expression Omnibus (GEO) database. Differentially expressed microRNAs (DEMs) were identified in BPD patients compared to control subjects, and their target genes were predicted using miRWalk, miRNet, miRDB, and TargetScan databases. Subsequently, protein-protein interaction (PPI) and functional enrichment analyses were conducted on the target genes. 30 hub genes were screened using the Cytohubba plugin of the Cytoscape software. Additionally, mRNA microarray data was utilized to validate the expression of hub genes and to perform immune infiltration analysis. Finally, real-time PCR (RT-PCR), immunohistochemistry (IHC), and flow cytometry were conducted using a mouse model of BPD to confirm the bioinformatics findings. Results: Two DEMs (miR-15b-5p and miR-20a-5p) targeting genes primarily involved in the regulation of cell cycle phase transition, ubiquitin ligase complex, protein serine/threonine kinase activity, and MAPK signaling pathway were identified. APP and four autophagy-related genes (DLC1, PARP1, NLRC4, and NRG1) were differentially expressed in the mRNA microarray dataset. Analysis of immune infiltration revealed significant differences in levels of neutrophils and naive B cells between BPD patients and control subjects. RT-PCR and IHC confirmed reduced expression of APP in a mouse model of BPD. Although the proportion of total neutrophils did not change appreciably, the activation of neutrophils, marked by loss of CD62L, was significantly increased in BPD mice. Conclusion: Downregulation of APP mediated by miR-15b-5p and miR-20a-5p may be associated with the development of BPD. Additionally, increased CD62L- neutrophil subset might be important for the immune-mediated injury in BPD.

Perinatal blockade of neuronal glutamine transport sex-differentially alters glutamatergic synaptic transmission and organization of neurons in the ventrolateral ventral media hypothalamus of adult rats.

Compared to male pups, perinatal female rats rely heavily on neuronal glutamine (Gln) transport for sustaining glutamatergic synaptic release in neurons of the ventrolateral ventral media nucleus of the hypothalamus (vlVMH). VMH mainly regulates female sexual behavior and increases glutamate release of perinatal hypothalamic neurons, permanently enhances dendrite spine numbers and is associated with brain and behavioral defeminization. We hypothesized that perinatal interruption of neuronal Gln transport may alter the glutamatergic synaptic transmission during adulthood. Perinatal rats of both sexes received an intracerebroventricular injection of a neuronal Gln uptake blocker, alpha-(methylamino) isobutyric acid (MeAIB, 5 mM), and were raised until adulthood. Whole-cell voltage-clamp recordings of miniature excitatory postsynaptic currents (mEPSCs) and evoked EPSCs (eEPSCs) of vlVMH neurons in adult rats with the perinatal pretreatment were conducted and neuron morphology was subjected to post hoc examination. Perinatal MeAIB treatment sex-differentially increased mEPSC frequency in males, but decreased mEPSC amplitude and synaptic Glu release in females. The pretreatment sex-differentially decreased eEPSC amplitude in males but increased AMPA/NMDA current ratio in females, and changed the morphology of vlVMH neurons of adult rats to that of the opposite sex. Most alterations in the glutamatergic synaptic transmission resembled the changes occurring during MeAIB acute exposure in perinatal rats of both sexes. We conclude that perinatal blockade of neuronal Gln transport mediates changes via different presynaptic and postsynaptic mechanisms to induce sex-differential alterations of the glutamatergic synaptic transmission and organization of vlVMH neurons in adult rats. These changes may be permanent and associated with brain and behavior feminization and/or defeminization in rats.

Multi-omics revealed the effects of rumen-protected methionine on the nutrient profile of milk in dairy cows.

Cow's milk is a highly-nutritious dairy product part of human diet worldwide. Rumen-protected methionine (RPM) is widely used to improve lactation performance of dairy cows, but understanding of the effects of RPM on milk nutrients composition are still limited. In this study, twenty mid-lactating dairy cows were supplemented with 20 gm/day RPM for 8 weeks to investigate the responses of milk nutritional composition to RPM. Metabolomics was applied for analyzing milk metabolites and 16S rRNA gene sequencing was used for analysis of rumen microbial composition. Milk fat content and yield were significantly increased after RPM supplementation. Totally 443 compounds belonging to 15 classes were identified, among which 15 metabolites were significantly changed. The functional nutrient α-ketoglutaric acid were significantly increased in the milk after RPM supplementation. We found 48 significantly differing bacterial genera in the rumen after supplementing RPM. Multi-omics integrated analysis revealed the higher abundance of Acetobacter, unclassified_f_Lachnospiraceae and Saccharofermentan contributed to the improved milk fat. In addition, the enriched abundance of Thermoactinomyces, Asteroleplasma, and Saccharofermentan showed positive correlations with higher α-ketoglutaric acid of milk. Our results uncover the metabolomic fingerprint and the key functional metabolites in the milk after supplementing RPM in dairy cows, as well as the key rumen bacteria associated with them. These findings provide novel insights into the development of functional dairy products that enriched the functional nutrient α-ketoglutaric acid or high milk fat.

Plastome variation and phylogeny of Taxillus (Loranthaceae).

Several molecular phylogenetic studies of the mistletoe family Loranthaceae have been published such that now the general pattern of relationships among the genera and their biogeographic histories are understood. Less is known about species relationships in the larger (> 10 species) genera. This study examines the taxonomically difficult genus Taxillus composed of 35-40 Asian species. The goal was to explore the genetic diversity present in Taxillus plastomes, locate genetically variable hotspots, and test these for their utility as potential DNA barcodes. Using genome skimming, complete plastomes, as well as nuclear and mitochondrial rDNA sequences, were newly generated for eight species. The plastome sequences were used in conjunction with seven publicly available Taxillus sequences and three sequences of Scurrula, a close generic relative. The Taxillus plastomes ranged from 121 to 123 kbp and encoded 90-93 plastid genes. In addition to all of the NADH dehydrogenase complex genes, four ribosomal genes, infA and four intron-containing tRNA genes were lost or pseudogenized in all of the Taxillus and Scurrula plastomes. The topologies of the plastome, mitochondrial rDNA and nuclear rDNA trees were generally congruent, though with discordance at the position of T. chinensis. Several variable regions in the plastomes were identified that have sufficient numbers of parsimony informative sites as to recover the major clades seen in the complete plastome tree. Instead of generating complete plastome sequences, our study showed that accD alone or the concatenation of accD and rbcL can be used in future studies to facilitate identification of Taxillus samples and to generate a molecular phylogeny with robust sampling within the genus.

Immobilized Co2+ and Cu2+ induced structural change of layered double hydroxide for efficient heterogeneous degradation of antibiotic.

In this study, MgMn-layered double hydroxide (MgMnLDH) exhibited excellent remediation functionality for heavy metals-antibiotics combined pollution. On the one hand, Co2+ and Cu2+ was efficiently immobilized on MgMnLDH with maximum quantity of 4.30 and 10.65 mmol g-1, respectively. A series of characterizations reflected the changes in structure and physicochemical properties of MgMnLDH after the immobilization. Density functional theory calculations (DFT) confirmed that the binding modes were lattice substitution for Co2+ and surface precipitation for Cu2+. On the other hand, the immobilized heavy metals enhanced the heterogeneous degradation for sulfamethoxazole (SMX) by peroxymonosulfate (PMS) activation. Complete degradation was achieved within 10 min in MgMnLDH-Co-4/PMS system and 60 min in MgMnLDH-Cu/PMS system, while only 20% in MgMnLDH/PMS system. The pH adaptability, reusability, stability and activation mechanism of two systems were systematically compared. The superior degradation performance of MgMnLDH-Co-4 benefited from the intense Co/Mn synergism and abundant oxygen vacancies, which could accelerate electron transfer during PMS activation process. The applicability of two catalysis system was confirmed in purifying other antibiotics and actual wastewater. The results highlight the importance of structural control in heterogeneous catalysis and provide new idea for environmental remediation.

Application of traditional clinical pathology quality control techniques to molecular pathology.

Many molecular diagnostic laboratories have evolved from research laboratories, initially performing low numbers of homebrew assays, but many laboratories now perform more kit-based assays, with ever increasing test volumes. One such assay is assessment of bone marrow transplantation engraftment. Allogeneic bone marrow transplantation is performed primarily in the treatment of hematological malignancies. Monitoring of engraftment was traditionally evaluated using minisatellites (variable number tandem repeats) and Southern blotting, but most laboratories now use Food and Drug Administration-cleared microsatellite (short tandem repeats) kits to assess the extent of engraftment. With the increase in equipment reliability, the use of kit-based assays, and the desire to provide the highest quality clinical data, we began applying traditional clinical pathology quality control tools to the molecular diagnostics laboratory. In this study, we demonstrate this approach using a microsatellite-based bone marrow engraftment assay. We analyzed control samples (pure and mixed) for two different microsatellites to establish quality control parameters and constructed Levey-Jennings charts to monitor both the precision and accuracy of this assay. By incorporating these tools into an overall quality assurance program, a laboratory can identify systematic errors and perform corrective actions before actual assay failure, thereby improving the quality of patient care.

Comparison of the CEDIA and MEIA assays for measurement of tacrolimus in organ transplant recipients.

OBJECTIVES: Tacrolimus is widely used in organ transplantation. We evaluated 2 immunoassays, CEDIA and MEIA, for the measurement of whole blood tacrolimus concentrations. METHODS: For each assay the following were evaluated: total precision, limit of detection (analytical sensitivity), limit of quantitation (functional sensitivity), linearity, and accuracy. Patient correlation studies were performed, comparing each assay with liquid chromatography-tandem mass spectrometry (LC-MS/MS). RESULTS: Total precision for MEIA, corresponding to mean concentrations of 6.8 and 22.5 ng/ml, was 17.4 and 11.9%, respectively. The limit of detection was 0.9 ng/ml, and the limit of quantitation was 4.7 ng/ml. Total precision for CEDIA, corresponding to mean concentrations of 5.3 and 19.9 ng/ml, was 20.6 and 6.3%, respectively. The limit of detection was 0.8 ng/ml, with a limit of quantitation of 4.9 ng/ml. Analysis of proficiency material demonstrated acceptable performance for both assays. In addition, both assays were acceptably linear over their respective reportable ranges. Analysis of patient correlation data using Passing-Bablok analysis and Bland-Altman plots demonstrated a positive average bias for both assays versus LC-MS/MS results. CONCLUSION: Based on our evaluation, both assays demonstrated acceptable performance for use in clinical monitoring of tacrolimus.

The glutamine-glutamate cycle regulates synaptic glutamate release in the ventrolateral ventromedial nucleus of the hypothalamus of perinatal female rats.

The astrocytic glutamine (Gln)-glutamate (Glu) cycle (GGC) supplies Gln for the regulation of glutamatergic synaptic transmission (GST) in the adult hippocampus. Increased synaptic Glu release in the perinatal ventrolateral ventromedial nucleus of the hypothalamus (vlVMH) modulates sexual differentiation, however, whether GGC regulates GST in the perinatal vlVMH has not been determined. Sex differences in oestradiol (E2 ) levels exist in the neonatal hypothalamus, and E2 increases levels of glutamine synthetase and glutaminase, two key enzymes involved in the GGC. Thus, it is hypothesised that sexually dimorphic phenotypes may exist in glutamatergic synapses associated with the GGC in the vlVMH in perinatal rats. Whole-cell voltage-clamp recordings in vlVMH neurones in brain slices from male and female pups revealed that pharmacological disruption of the GGC by α-(methylamino) isobutyric acid (5 mmol L-1 ), which blocks neuronal Gln uptake; or by l-methionine sulphoximine (1.5 mmol L-1 ), which inhibits astrocytic Gln synthesis, decreased miniature excitatory postsynaptic current (mEPSC) amplitudes in female but not male pups. By contrast, GGC interruptions decreased evoked (e)EPSC amplitudes in both sexes following increased synaptic activity produced by a period of stimulation. In male pups, the decreased eEPSCs were attributable to reduced Glu release, as assessed by paired-pulse stimulations, whereas, in female pups, they were attributable to decreased Glu content in the synaptic vesicles, as measured by strontium-evoked mEPSCs. The l-methionine sulphoximine-mediated decrease in eEPSCs was rapidly rescued by exogenous Gln in female but not male pups. The reductions in mEPSCs and eEPSCs in female pups were accompanied by enhanced blocking effects of the low-affinity Glu AMPA receptor antagonist, γ-d-glutamylglycine, consistent with diminished Glu release. In conclusion, female, but not male pups, rely on constitutive astrocytic Gln for sustained synaptic Glu release in the vlVMH. This glutamatergic synaptic phenotype may be associated with brain and behaviour feminisation and/or defeminisation in rats.

Dynamic regulation of synaptic GABA release by the glutamate-glutamine cycle in hippocampal area CA1.

Vesicular GABA and intraterminal glutamate concentrations are in equilibrium, suggesting inhibitory efficacy may depend on glutamate availability. Two main intraterminal glutamate sources are uptake by neuronal glutamate transporters and glutamine synthesized through the astrocytic glutamate-glutamine cycle. We examined the involvement of the glutamate-glutamine cycle in modulating GABAergic synaptic efficacy. In the absence of neuronal activity, disruption of the glutamate-glutamine cycle by blockade of neuronal glutamine transport with alpha-(methylamino) isobutyric acid (MeAIB; 5 mM) or inhibition of glutamine synthesis in astrocytes with methionine sulfoximine (MSO; 1.5 mM) had no effect on miniature IPSCs recorded in hippocampal area CA1 pyramidal neurons. However, after a period of moderate synaptic activity, application of MeAIB, MSO, or dihydrokainate (250 microM; an astrocytic glutamate transporter inhibitor) significantly reduced evoked IPSC (eIPSC) amplitudes. The MSO effect could be reversed by exogenous application of glutamine (5 mM), whereas glutamine could not rescue the eIPSC decreases induced by the neuronal glutamine transporter inhibitor MeAIB. The activity-dependent reduction in eIPSCs by glutamate-glutamine cycle blockers was accompanied by an enhanced blocking effect of the low-affinity GABA(A) receptor antagonist, TPMPA [1,2,5,6-tetrahydropyridin-4-yl)methylphosphinic acid], consistent with diminished GABA release. We further corroborated this hypothesis by examining MeAIB effects on minimal stimulation-evoked quantal IPSCs (meIPSCs). We found that, in MeAIB-containing medium, moderate stimulation induced depression in potency of meIPSCs but no change in release probability, consistent with reduced vesicular GABA content. We conclude that the glutamate-glutamine cycle is a major contributor to synaptic GABA release under physiological conditions, which dynamically regulates inhibitory synaptic strength.

Tumour suppressor function of RNase L in a mouse model.

RNase L is one of the key enzymes involved in the molecular mechanisms of interferon (IFN) actions. Upon binding with its activator, 5'-phosphorylated, 2'-5' oligoadenylates (2-5A), RNase L plays an important role in the antiviral and anti-proliferative functions of IFN, and exerts proapoptotic activity independent of IFN. In this study, we have found that RNase L retards proliferation in an IFN-dependent and independent fashion. To directly measure the effect of RNase L on tumour growth in the absence of other IFN-induced proteins, human RNase L cDNA was stably expressed in P-57 cells, an aggressive mouse fibrosarcoma cell line. Three clonal cell lines were isolated in which the overexpression of RNase L was 15-20-fold of the endogenous level. Groups of five nude mice were injected subcutaneously with either the human RNase L overexpressing clones (P-RL) or control cells transfected with an empty vector (P-Vec). Tumour growth by the two cell lines was monitored by measuring tumour volumes. In the P-RL group, tumour formation was significantly delayed and the tumours grew much slower compared to the control group. Morphologically, the P-RL tumour appeared to have more polygonal cells and increased single cell tumour necrosis. Interestingly, P-RL tumours eventually started to grow. Further analysis revealed, however, that these tumours no longer expressed ectopic RNase L. Our findings suggest that RNase L plays a critical role in the inhibition of fibrosarcoma growth in nude mice.

Enzymes and related proteins as cancer biomarkers: a proteomic approach.

BACKGROUND: The discovery of cancer biomarkers has become a major focus of cancer research, which holds promising future for early detection, diagnosis, monitoring disease recurrence and therapeutic treatment efficacy to improve long-term survival of cancer patients. Most of the functional information of the cancer-associated genes resides in the proteome. Since cancer is a complex disease, it might require a panel of multiple biomarkers in order to achieve sufficient clinical efficacy. METHODS: Serum/plasma is the most accessible biological specimen collected from patients. Therefore, serum proteomic diagnostics would be the most promising new test for cancer. With the advent of new and improved proteomic technologies, such as protein chips and mass spectrometry coupled with advanced bioinformatic tools, it is possible to develop potential cancer biomarkers. However, specimen collection, handling, study design and data analysis are essential components for successful biomarker discovery and validation. Multi-center case control study should be conducted with extensive clinical validation to minimize the impact of possible confounding variables (non-biological). CONCLUSIONS: Enzymes and related proteins, such as inhibitors, are promising candidates for cancer diagnostics.

Genetic, antigenic, and pathogenic characteristics of avian infectious bronchitis viruses genotypically related to 793/B in China.

In this study, 20 infectious bronchitis virus (IBV) strains, which were genotypically related to 793/B, as assessed by an S1 gene comparison and a complete genomic sequence analysis, were isolated and identified from 2009 to 2014 in China. Phylogenetic analysis, network tree, similarity plot analysis, Recombination Detection Program 4(RDP4) and sequence comparison revealed that 12 of the 20 isolates were likely the reisolated vaccine virus. One isolate, ck/CH/LSD/110857, was shown to have originated from recombination events between H120- and 4/91-like vaccine strains that did not result in changes of antigenicity and pathogenicity. The remaining seven IBV isolates were shown to have originated from recombination events between a 4/91-like vaccine strain and a GX-LY9-like virus, which were responsible for the emergence of a novel serotype. A vaccination-challenge test found that vaccination with the 4/91 vaccine strain did not provide protection against challenge with the recombinant viruses. In addition, the results showed that the recombination events between the vaccine and field strains resulted in altered genetics, serotype, antigenicity, and pathogenicity compared with those of their deduced parental viruses. The results are important not only because this virus is of economic importance to poultry industry, but also because it is important for elucidating the origin and evolution of other coronaviruses.

Origin and evolution of LX4 genotype infectious bronchitis coronavirus in China.

We investigated the genomic characteristics of 110 LX4 genotype strains of infectious bronchitis viruses (IBVs) isolated between 1995 and 2005 in China. The genome of these IBVs varies in size from 27596bp to 27790bp. Most IBV strains have the typical genomic organization of other gamacoronaviruses, however, two strains lacked 3a and 5b genes as a result of a nucleotide change within the start codon in the 3a or 5b genes. Analysis of our 110 viruses revealed that recombination events may be responsible for the emergence of the LX4 genotype with different topologies. Most of these viruses disappeared (before mid-2005) because they were not "fit" to adaptation in chickens. Finally, those of the "fit" viruses (after mid-2005) continued to evolve and have become widespread and predominant in commercial poultry. In addition, few of these viruses experienced recombination with those of the vaccine strains at the 3' end of the genome.

Clioquinol effects on tissue chelatable zinc in mice.

Recent evidence for the involvement of zinc in the formation of beta-amyloid plaques in the brain in Alzheimer's disease has led to the establishment of new therapeutic strategies for the degenerative disorder based on metal chelation. The present experiment was conducted on a membrane-permeable zinc chelator, clioquinol (CQ), that has shown potential in initial studies on a mouse model of Alzheimer's disease [1]. The degree of chelatable zinc in mice treated with CQ, delivered by two different routes, was measured using complementary protocols for identifying chelatable zinc: 6-methoxy-8-quinolyl- p-toluenesulfonamide (TSQ) histofluorescence, and selenite autometalography. Mice injected intraperitoneally with CQ showed a dramatic reduction in chelatable zinc in brain, testis, and pancreas. In contrast, mice given CQ orally showed no significant change in levels of chelatable zinc in these tissues. This suggests that CQ administered orally to patients with Alzheimer's disease should not significantly perturb chelatable zinc levels in key organs and may be used over long periods without adverse endocrinological and reproductive effects related to zinc deficiency. In contrast, CQ injected intraperitoneally may be used not only as a tool for investigating chelatable zinc pools but also in a clinical context. For example, injected CQ could be employed in situations requiring the rapid buffering of excessive chelatable zinc following ischemic episodes or brain trauma. Thus, our findings indicate that CQ has considerable potential as a versatile scientific and clinical tool used for selective modulation of zinc pools.