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Saudi Arabia has an overall smoking rate of 15.9%. The link between smoking and periodontal disease has been studied extensively. It is possible for human gingival fibroblasts to accumulate nicotine intracellularly over a period of four hours. Additionally, unmetabolized nicotine is released into the environment. Tobacco presence can impair tissue inflammation, wound healing, and organ development. To counterbalance tobacco toxins, vitamin C has been added to a variety of products. This study aims to analyze the RNA expression of antioxidant, anti-inflammatory, and wound healing proteins in human gingival fibroblasts from smokers and nonsmokers using polymerase chain reaction. hGFs were extracted from clinically healthy periodontium sites of adult male subjects. Both heavy cigarette smokers and never-smokers participated as subjects. Cells were cultured and subcultured in supplemented growth medium. Vitamin C was inducted in the medium at the experimental 6th passage. RNA expression analysis (qRT-PCR) was performed to analyze adhesion, proliferation, and extracellular matrix expression. The results revealed marked expression of a wound healing gene (VEGF-A) in never-smokers (p value = 0.016). GPX3 and SOD3 represent antioxidants that are highly expressed in treated never-smoker cells. SOD2 significantly increased (p value = 0.016) in smokers after vitamin C exposure. The anti-inflammatory markers IL-6 and IL-8 were lower among smokers than among nonsmokers (p < 0.0001). Tobacco smoking suppressed gingival fibroblasts' abilities to regenerate, heal, combat inflammation, and resist free radicals. Vitamin C at cellular levels was beneficial and should be considered in the treatment component of smokers in the dental clinic. [ABSTRACT FROM AUTHOR]

Lung adenocarcinoma (LUAD), the predominant subtype of lung cancer, has a high incidence and annual mortality worldwide. Members of the cadherin-related (CDHR) family are associated with many malignant tumor types. However, their role and clinical significance in LUAD have not been clarified. We analyzed the association of CDHRs mRNA expression profiles with prognostic significance, immune infiltration, and potential biological functional signatures in several public databases. We constructed a co-expressed mRNA network, and performed an intrinsic molecular structure and function enrichment analysis. Our results showed that CDHR2 mRNA expression was upregulated in LUAD, whereas CDHR1, CDHR3, CDHR4, and CDHR5 mRNAs were downregulated. Upregulation of CDHR2 mRNA is associated with a poor prognosis in patients with LUAD. Next, the correlation between CDHR family members and immune infiltration was observed. A receiver operating characteristic curve showed that the CDHR family is valuable for diagnosing LUAD. In this study, we found that CDHR2 mRNA expression was upregulated in LUAD. Upregulation of CDHR2 mRNA was associated with a poor prognosis in patients with LUAD. [ABSTRACT FROM AUTHOR]

Osteoarthritis (OA) is a widespread joint disorder that is primarily noted for the progressive degeneration of joint cartilage, accompanied by a significant inflammatory response. Recently, there has been a growing interest in understanding the roles of ATF3 and ferritin-related RNAs in the context of immune responses and inflammatory processes. However, their specific functions and mechanisms in the progression of osteoarthritis have remained largely ambiguous and underexplored. The primary objective of this study was to thoroughly investigate the changes in expression levels of ATF3 and ferritin-related RNAs within osteoarthritic tissues, as well as to examine their potential effects on immune cell infiltration. To achieve this, advanced RNA sequencing technology was employed to meticulously analyze the expression levels of ATF3 and the ferritin-related RNAs. Furthermore, bioinformatics methods were utilized to assess the infiltration patterns of various immune cells and to explore the correlation between these infiltration patterns and the expression levels of RNA. The findings from this study revealed that both ATF3 and ferritin-related RNAs exhibited significantly elevated expression levels in tissues affected by osteoarthritis. Additionally, the immunoinfiltration analysis highlighted a positive correlation between the degree of infiltration of T cells and macrophages and the levels of ferritin-related RNAs. Such findings suggest that the presence of these immune cells is intricately linked to the expression of ferritin-associated RNAs. Further investigations indicated that ferritin-associated RNAs play a critical role in the progression of osteoarthritis by modulating inflammatory responses and influencing the activity of various immune cells. Consequently, both ATF3 and ferritin-related RNAs demonstrate abnormal expression patterns in osteoarthritis, which are closely associated with the infiltration of immune cells. [ABSTRACT FROM AUTHOR]

Smoking and the severity of periodontal disease have long been associated. In Saudi Arabia, tobacco smoking is rising, contributing to the increased demand for products that counter its detrimental effects. The antioxidant properties of vitamin C (vit C) make it a powerful countermeasure to tobacco toxicity. Observation of these effects on human gingival fibroblasts (hGFs) would suggest use of vitamin C in future dental applications. To examine the proliferation, adhesion, and expression of extracellular RNA in human gingival fibroblasts extracted from cigarette smokers when compared to never-smokers, in association with vitamin C. Human gingival fibroblasts were extracted from Periodontal free sites of healthy adult male participants. Group 1; consisted of Heavy cigarette smokers (n = 1) while group 2 was never-smokers (n = 1). Collected cells were cultured and subcultured in supplemented growth medium. Vitamin C was then induced in the medium at the experimental sixth passage. RNA expression analysis using quantitative reverse transcriptase-polymerase chain reaction was performed to analyze the adhesion, proliferation, and extracellular matrix expression. Expression of the adhesion gene (CD44) in the smoker group was significantly downregulated than never-smoker group (p-value = 0.024). After the induction of vitamin C, the smoker samples showed a significant improvement in their gene expression levels. The extracellular genes involved in this study (COL1A1 , LAMA3 , and TGFB3) were significantly affected by the smoking status. In addition, the proliferation of MK167 and CCNB1 genes in smokers and never-smokers was increased. Cigarette smoking affects the overall properties of human gingival fibroblasts' adhesion, proliferation, and extra-cellular matrix formation. Furthermore, the addition of vitamin C affects these cellular properties in a positive manner. [ABSTRACT FROM AUTHOR]

• Fermentations with SSL resulted in the upregulation of pepck and mdh that participate in the reductive TCA cycle. • Fumarate reductase that participates in oxidative phosphorylation and TCA cycle was significantly upregulated in SSL fermentations compared to pure substrates. • SSL and xylose resulted in similar and significant downregulation of ATP synthase compared to glucose. The fermentative production of biobased chemicals and polymers using crude lignocellulose hydrolysates is challenging due to the presence of various inhibitory compounds and multiple sugars. This study evaluates the metabolic response of Actinobacillus succinogenes for the production of succinic acid using spent sulphite liquor (SSL) as feedstock derived from industrial acidic sulphite pulping of Eucalyptus globulus hardwood. A transcriptomic approach led to significant insights on gene regulation of the major metabolic pathways (glycolysis, pentose phosphate pathway, TCA cycle, pyruvate metabolism and oxidative phosphorylation) in batch cultures carried out on SSL and compared with glucose and xylose. Significantly overexpressed genes in SSL compared to glucose and xylose were fructose biphosphate aldolase (> 1.18-fold change) in the catabolism, phosphoenolpyruvate carboxykinase (> 1.59-fold change) and malate dehydrogenase (> 1.49-fold change) in the TCA cycle, citrate lyase (> 1.7-fold change), dihydrolipoamide dehydrogenase (> 0.88-fold change), pyruvate dehydrogenase E2 (> 1.63-fold change) and pyruvate formate lyase (> 0.61-fold change), involved in acetyl-CoA pathways. Finally, C4 tricarboxylic transporters were overexpressed (DCU (> 1.61-fold change) and 0079 (> 4.19-fold change). SSL was responsible for the upregulation of genes involved in the TCA cycle and oxidative phosphorylation, while xylose showed similar results with SSL in the oxidative phosphorylation. [ABSTRACT FROM AUTHOR]

Renal oncocytoma (RO) accounts for 5% of renal cancers and generally behaves as a benign tumor with favorable long-term prognosis. It is difficult to confidently distinguish between benign RO and other renal malignancies, particularly chromophobe renal cell carcinoma (chRCC). Therefore, RO is often managed aggressively with surgery. We sought to identify molecular biomarkers to distinguish RO from chRCC and other malignant renal cancer mimics. In a 44-patient discovery cohort, we identified a significant differential abundance of nine genes in RO relative to chRCC. These genes were used to train a classifier to distinguish RO from chRCC in an independent 57-patient cohort. The trained classifier was then validated in five independent cohorts comprising 89 total patients. This nine-gene classifier trained on the basis of differential gene expression showed 93% sensitivity and 98% specificity for distinguishing RO from chRCC across the pooled validation cohorts, with a c-statistic of 0.978. This tool may be a useful adjunct to other diagnostic modalities to decrease the diagnostic and management uncertainty associated with small renal masses and to enable clinicians to recommend more confidently less aggressive management for some tumors. Renal oncocytoma is generally a benign form of kidney cancer that does not necessarily require surgical removal. However, it is difficult to distinguish renal oncocytoma from other more aggressive forms of kidney cancer, so it is treated most commonly with surgery. We built a classification tool based on the RNA levels of nine genes that may help avoid these surgeries by reliably distinguishing renal oncocytoma from other forms of kidney cancer. We trained and validated a classifier to distinguish renal oncocytoma from chromophobe renal cell carcinoma based on differential RNA expression. This classifier may be used to decrease diagnostic and management uncertainty associated with small renal masses. [ABSTRACT FROM AUTHOR]

Estuaries are thought to be very harsh environments because their physico-chemical parameters, such as salinity, temperature and oxygen, can fluctuate substantially. The distribution of species living in an estuary is largely determined by how well they can cope with such fluctuations. The spionid polychaete Pygospio elegans is common in boreal estuarine habitats that show strong salinity fluctuations, living in habitats ranging from fully marine salinities to brackish environments with salinities as low as 5. In this study we investigated the abilities of P. elegans to cope with an acute as well as a long-term change in salinity. Specimens originating from a salinity of about 15 were exposed to salinity 15 as control and 5 and 30 as low and high salinity treatments. In acute exposure experiments, we measured body volume and tissue water content as well as gene expression of seven genes related to volume- and osmoregulation within 4 h of exposure. In a long-term experiment, we monitored survival, growth and reproduction as well as potential changes in DNA methylation within 6 weeks after a gradual salinity change. In response to abrupt exposure to a hyposmotic medium (salinity 5) we observed increased body volume that could not be fully restored, increased mortality, and no clear change in gene expression. Hence, P. elegans might be a weak cell-volume regulator that cannot cope well with sudden drops in salinity. In contrast, a gradual, long-term change in salinity seemed to be less stressful, although worms at salinity 5 showed slightly increased mortality and reduced or delayed maturity. Overall, there were fewer detrimental effects of exposure to a higher salinity. While the tested salinities fall within the tolerance range of P. elegans from this population, decrease in salinity seems to be a stronger stress. • Pygospio elegans is a weak volume regulator under hyposmotic conditions. • RNA expression analyses showed no indication of osmoregulation. • Abrupt drops in salinity might affect population size and metapopulation dynamics. • Hyposmotic conditions slightly lower/delay growth and maturation in the long-term. [ABSTRACT FROM AUTHOR]

Amyotrophic lateral sclerosis (ALS) is a generally fatal neurodegenerative disease of adults that produces weakness and atrophy due to dysfunction and death of upper and lower motor neurons. We used RNA-sequencing (RNA-seq) to analyze expression of all mitochondrial DNA (mtDNA)-encoded respiratory genes in ALS and CTL human cervical spinal cords (hCSC) and isolated motor neurons. We analyzed with RNA-seq mtDNA gene expression in human neural stem cells (hNSC) exposed to recombinant human mitochondrial transcription factor A (rhTFAM), visualized in 3-dimensions clustered gene networks activated by rhTFAM, quantitated their interactions with other genes and determined their gene ontology (GO) families. RNA-seq and quantitative PCR (qPCR) analyses showed reduced mitochondrial gene expression in ALS hCSC and ALS motor neurons isolated by laser capture microdissection (LCM), and revealed that hNSC and CTL human cervical spinal cords were similar. Rats treated with i.v. rhTFAM showed a dose-response increase in brain respiration and an increase in spinal cord mitochondrial gene expression. Treatment of hNSC with rhTFAM increased expression of mtDNA-encoded respiratory genes and produced one major and several minor clusters of gene interactions. Gene ontology (GO) analysis of rhTFAM-stimulated gene clusters revealed enrichment in GO families involved in RNA and mRNA metabolism, suggesting mitochondrial-nuclear signaling. In postmortem ALS hCSC and LCM-isolated motor neurons we found reduced expression of mtDNA respiratory genes. In hNSC’s rhTFAM increased mtDNA gene expression and stimulated mRNA metabolism by unclear mechanisms. rhTFAM may be useful in improving bioenergetic function in ALS. [ABSTRACT FROM AUTHOR]

To improve our understanding of the abnormalities and non-Mendelian inheritance characteristics of schizophrenia, this study examined DNA methylation (5mC) and hydroxymethylation (5hmC) in the schizophrenia-associated GABRB2 gene encoding the type A γ-aminobutyric acid receptor β 2 subunit. DNAs from the peripheral white blood cells of 279 schizophrenic patients and 256 controls from the Chinese Han population were examined to reveal that the GABRB2 promoter P1-5mC level which was correlated with olanzapine administration, P2-5mC/5hmC level, and Alu-5mC level which was correlated with administration of ziprasidone or oxcarbazepine, were increased in schizophrenic patients. Significant correlations of the promoter 5mC/5hmC levels with the genotypes of single nucleotide polymorphisms (SNPs) were observed with SNPs rs72815526 (C/A) and rs3811997 (C/T). In schizophrenics, the heterozygous genotypes of rs72815526 (C/A) were correlated with increased 5hmC levels whereas the heterozygous genotypes of rs3811997 (C/T) were correlated with decreased 5mC levels. Moreover, the GABRB2 promoter with rs3811997(C/T) minor allele T or the methylation-disrupted type AG at −254 and −231 CCGG sites was observed to enhance the promoter activity in the luciferase reporter-transfected human embryonic kidney 293 cells. An elevated GABRB2 mRNA transcriptional level in human neuroblastoma IMR32 cells were accompanied by the decreased promoter 5hmC/5mC levels induced by 5-azacytidine or by increased histone H4 acetylation levels of the Alu-Yi6 region induced by valproic acid. These results reveal alterations in GABRB2 genotype-dependent methylation and hydroxymethylation in schizophrenia, which yielded transcriptional and translational alterations in the cultured cells, and help elucidate the genetic-epigenetic interactions influencing schizophrenia. [ABSTRACT FROM AUTHOR]

• Increased succinate production via microbial electrosynthesis with A. succinogenes. • FRD and cytochromes of oxidative phosphorylation were downregulated. • PEPCK that participates in rTCA cycle was upregulated. The potential of renewable energy application via direct electrode interaction for the production of bio-based chemicals is a promising technology. The utilization of extracellular energy in pure culture fermentations aims in intracellular redox balance regulation in order to improve fermentation efficiency. This work evaluates the impact of a bioelectrochemical system in succinic acid fermentation and the metabolic response of Actinobacillus succinogenes. The metabolic pathway regulation of A. succinogenes was evaluated via RNA expression of the key enzymes that participate in TCA cycle, pyruvate metabolism and oxidative phosphorylation. The genes that were significantly overexpressed in BES compared to non-BES were phosphoenolpyruvate carboxykinase (0.4-fold change), inorganic pyrophosphatase (2.3-fold change) and hydrogenase (2.2-fold change) and the genes that were significantly underexpressed were fumarase (−0.94-fold change), pyruvate kinase (−6.9-fold change), all subunits of fumarate reductase (−2.1 to −1.17-fold change), cytochromes I and II (−1.25 and −1.02-fold change, respectively) and two C4-carboxylic acid transporters. [ABSTRACT FROM AUTHOR]

Niu Huang Jie Du prescription (NHJD) is a traditional Chinese medicine (TCM) widely used in patients suffering from excessive inner fire toxin (Huo Du Nei Sheng) syndrome, such as sore throat, gingival swelling, and pain, mouth and tongue sores, etc. This formula contains realgar (As 4 S 4) which is one of the 28 toxic medicinal materials promulgated by the Chinese Ministry of Health. Many studies reported its toxicity on the liver and kidney, and the detoxification effect of NHJD. However, its detoxification mechanism is still unclear. To clarify the detoxification mechanism of NHJD to realgar, this study evaluated the detoxification effect of NHJD on realgar exposure in mice, and analyzed differences in mRNA expression profiles in liver tissues and associated functional predictions. ICR mice were administered with NHJD, realgar, and CMC-Na as blank control for 12 weeks, respectively. Liver injury was evaluated by histopathologic examination and liver mRNA gene were sequenced by Illumina. Differentially expressed gene, functionally enrichment and protein association network analysis were conducted. 43 genes were screened out, among which 15 genes in the realgar group were decreased, but the extent of the decline has been restored in the NHJD group. The remaining 28 genes have exactly the opposite trends. Functional module analysis revealed that those detoxification function-related genes were primarily for positive regulation of glutathione metabolism, P450 on the metabolism of exogenous compounds, oxidative stress and immune-related, etc. The results indicated that realgar mainly causes liver damage by changing the common enzymes of drug metabolism, especially the expression of genes related to CYPs, GSTs family, oxidative stress, and complement immunity, while the TCM prescription NHJD has a regulatory effect on the abnormal expression of corresponding genes. Our results will provide some clues for the detoxification mechanism of arsenic-containing TCM prescriptions. [Display omitted] • We focused on the detoxification mechanism of realgar-containing NHJD in mice. • RNA expression, KEGG, GO analysis and protein association network were conducted. • Enzymes of drug metabolism may be involved in the detoxification mechanism of NHJD. [ABSTRACT FROM AUTHOR]

Fluorescent in situ hybridization (FISH) is a powerful technique for the detection of RNA or DNA within cells and tissues, which provides a unique link between molecular and cell biology. This technique is broadly applicable across a range of biological systems. While FISH has been previously adapted to flow-based platforms, their use remains limited because of procedural challenges and costs associated with commercial kits. Herein we present a protocol that modifies existing techniques to sensitively and specifically detect and examine RNA expression patterns in primary cells and cell lines using flow cytometry (expression-FISH; X-FISH). As relevant examples, we show how this technique can be used to monitor changes in mRNA expression following activation, how it can be combined with antibody staining to study RNA and protein in the same sample, and how it can help distinguish among subsets in a mixed cell population. X-FISH can integrate multiple probes and can be performed in conjunction with other assays, allowing for informative multiparametric analyses and increased statistical robustness. For non-classical comparative animal models this procedure provides a time saving alternative to de novo production of antibody-based markers. Finally, X-FISH provides an economical solution that is applicable to conventional as well as multi-spectral imaging flow cytometry platforms. [ABSTRACT FROM AUTHOR]

Background Oxytocin is a neuropeptide that is involved in the regulation of mood, anxiety and social biology. Genetic variation in the oxytocin receptor gene (OXTR) has been implicated in anxiety, depression and related stress phenotypes. It is not yet known whether OXTR interacts with other risk factors such as early life trauma to heighten the severity of experienced anxiety and depression. Methods In this study, we examined genotypes in 653 individuals and tested whether SNP variation in OXTR correlates with severity of features of self-reported experience on the Depression Anxiety and Stress Scale (DASS), and whether this correlation is enhanced when early life trauma is taken into account. We also assessed the effects of OXTR SNPs on RNA expression levels in two separate brain tissue cohorts totaling 365 samples. Results A significant effect of OXTR genotype on DASS anxiety, stress and depression scores was found and ELS events, in combination with several different OXTR SNPs, were significantly associated with differences in DASS scores with one SNP (rs139832701) showing significant association or a trend towards association for all three measures. Several OXTR SNPs were correlated with alterations in OXTR RNA expression and rs3831817 replicated across both sets of tissues. Conclusions These results support the hypothesis that the oxytocin system plays a role in the pathophysiology of mood and anxiety disorders. [ABSTRACT FROM AUTHOR]

Understanding miRNAs' regulatory networks and target genes could facilitate the development of therapies for human diseases such as cancer. Although much useful gene expression profiling data for tumor cell lines is available, microarray data for miRNAs and mRNAs in the human HepG2 cell line have only been compared with that of other cell lines separately. The relationship between miRNAs and mRNAs in integrated expression profiles for HepG2 cells is still unknown. To explore the miRNA-mRNA correlations in hepatocellular carcinoma (HCC) cells, we performed miRNA and mRNA expression profiling in HepG2 cells and normal liver HL-7702 cells at the genome scale using next-generation sequencing technology. We identified 193 miRNAs that are differentially expressed in these two cell lines. Of these, 89 miRNAs were down-regulated in HepG2 cells compared with HL-7702 cells, while 104 miRNAs were up-regulated. We also observed 3035 mRNAs that are significantly dys-regulated in HepG2 cells. We then performed an integrated analysis of the expression data for differentially expressed miRNAs and mRNAs and found several miRNA-mRNA pairs that are significantly correlated in HepG2 cells. Further analysis suggested that these differentially expressed genes were enriched in four tumorigenesis-related signaling pathways, namely, ErbB, JAK-STAT, mTOR, and WNT, which until now had not been fully reported. Our results could be helpful in understanding the mechanisms of HCC occurrence and development. [ABSTRACT FROM AUTHOR]

Abstract: Using two microarray platforms, we identify HLA-DRB5 as the most highly expressed gene in MS compared to healthy subjects. As expected, HLA-DRB5 expression was associated with the HLA-DRB1*1501 MS susceptibility allele. Besides HLA-DRB5, there were 1219 differentially expressed exons (p<0.01, |fold change (FC)|>1.2) that differed between HLA-DRB1*1501 Positive multiple sclerosis subjects (MSP) compared to HLA-DRB1*1501 negative multiple sclerosis subjects (MSN). Analysis of the regulated genes revealed significantly different immune signaling pathways including IL-4 and IL-17 in these two MS genotypes. Different risk alleles appear to be associated with different patterns of gene expression that may reflect differences in pathophysiology of these two MS subtypes. These preliminary data will need to be confirmed in future studies. [Copyright &y& Elsevier]

Abstract: Fixation techniques preserving morphological fidelity, protein antigenicity and integrity of nucleic acids can have a high impact on both basic and applied biomedical sciences and diagnostic pathology. Different types of mouse tissues were fixed with neutral buffered formalin, ethanol supplemented with acetic acid and modified methacarn (methanol-Carnoy) fixative. The alcohol-fixed samples were processed in an Autotechnicon tissue processor or in an incubator. The preservation of tissue morphology was assessed in all specimens and the immunoreactivity was evaluated with antibodies specific for proteins with nuclear, membrane or cytoplasmic localization. RNA was extracted from all groups of fixed hind limb skeletal muscle specimens and was assessed versus unfixed tissue for preservation of its quantity and quality by amplification of gene-specific fragments of different lengths. Both alcohol-based fixatives preserved the tissue architecture and the specificity of immunoreactivity in excellent quality; the trimming approach did not result in detectable differences. Oligonucleotide fragments of length between 108 and 577 base pairs were amplified from all groups of alcohol-fixed skeletal muscle specimens in amounts comparative to the unfixed muscle tissue. We conclude that both alcohol-based fixatives are an excellent tool for storage of tissue samples designed for immunohistochemical and mRNA expression studies when the access to fresh samples is limited. [Copyright &y& Elsevier]

Abstract: Introduction: Deep hypothermic circulatory arrest (DHCA) is used in corrective cardiac surgery for complex congenital heart disease. Endogenous protective mechanisms may be responsible for the prevention of brain damage after hypothermic ischemia. Neuroglobin and cytoglobin are expressed in brain cells and appear to modulate hypoxic-ischemic brain injury. However, their neuroprotective potency is still not understood. Thus the aim of this study was to detect the influence exerted by DHCA on their expression. Methods: The effects of DHCA were analyzed in a neonatal piglet model with cardiopulmonary bypass, DHCA of 60 and 120min and subsequent reperfusion of 6h. Complete histological analysis and changes in the mRNA expression of neuroglobin and cytoglobin were measured in the brain. Results: In comparison to animals without DHCA, neuroglobin expression was stable after 60min DHCA and neuronal cell necrosis in the cortex was mild (< 10 %). Neuroglobin expression was significantly reduced after 120min DHCA, which was accompanied by substantial neuronal cell necrosis (> 50 %). Cytoglobin expression did not differ significantly between animals with neuronal necrosis vs. sham. Conclusion: Constitutive expression levels of neuroglobin may explain the mild neuronal injury after 60min DHCA. Significant neuronal cell death correlates with reduced neuroglobin expression and might reflect a limited capacity to compensate for ischemic injury. Both respiratory cell proteins may constitute attractive targets for therapeutic modulation of gene regulation, but further studies are necessary. [ABSTRACT FROM AUTHOR]

Abstract: Placental alkaline phosphatase (PLAP), encoded by the ALPP gene, is produced by the fetal side of the placenta. This enzyme displays strong genetic variability. Some of the variants were reported to be associated with pathology of pregnancy. We show here that the two most common ALPP allelic variants, Pl1 and Pl2 , differ in mRNA expression level. This differential expression was independent of the parental origin and probably results from linkage disequilibrium with the sequence variation rs2014683G>A in the ALPP gene promoter that was shown to have allele-specific binding patterns to placental nuclear proteins. The possible role of this allelic-specific expression in placenta-related pathology is discussed. [Copyright &y& Elsevier]

Abstract: Apelin is a recently discovered peptide produced by several tissues including brain and adipose tissue. In mammals and zebrafish, apelin regulates cardiovascular functions. Recent evidence in mammals suggest that apelin might also regulate food intake. In this study, we cloned a cDNA encoding apelin and examined apelin mRNA distribution within the brain and in peripheral tissues. We also assessed the effects of fasting on apelin brain mRNA abundance. Apelin mRNA was expressed throughout the brain as well as in several peripheral tissues including brain, spleen, heart and fat. Apelin mRNA abundance in both hypothalamus and telencephalon was significant higher in fasted fish than in fed fish. In order to further characterize apelin in goldfish, we assessed the effects of central (intracerebroventricular, icv) and peripheral (intraperitoneal, ip) injections of apelin-13 on food intake in goldfish. Apelin injected ip at a dose of 100ng/g or icv at a dose of 10ng/g induced a significant increase in food intake compared to saline-injected fish. Our results suggest that apelin acts as an orexigenic factor in goldfish. Its widespread distribution in the brain and the periphery also suggests that apelin might have multiple physiological regulating roles in fish. [Copyright &y& Elsevier]

Malignant pleural mesothelioma is an aggressive malignancy for which surgical treatment is an option for a subset of patients. A gene ratio test based on the relative expression levels of four genes has been validated in clinical trials to predict which patients will benefit from surgical therapy. A description of the test properties and the independent predictors for outcome are provided. [Copyright &y& Elsevier]

Although irradiation has been used to destroy bacteria and molds in spices globally, a new technology is needed to alleviate public fears of irradiation, the high cost of irradiated food, and environmental contamination at the irradiation facility. The objective of this study was to investigate the effects of dielectric barrier discharge (DBD) plasma on Eshcericah coli O157: H7 and Bacillus cereus levels in red pepper powder and the physicochemical properties of red pepper powder stored at 25 °C. RNA expression was determined by qRT-PCR. The surface of bacteria treated with DBD plasma was observed with a Field Emission Scanning Electron Microscope (FE-SEM). Red pepper powder (5 g) inoculated with E. coli O157: H7 and B. cereus was treated with DBD plasma using argon as a plasma-forming gas at 31 kW for 0, 5, 10, and 15 min. DBD plasma did not significantly decrease the bacterial concentrations with an increase in treatment time. However, DBD plasma treatment for 15 min killed E. coli O157: H7 more rapidly during storage. Expression of virulence genes of both E. coli O157: H7 and B. cereus was decreased by DBD plasma treatment for 15 min. Results of FE-SEM showed that DBD plasma damaged the bacterial cell membranes, leading to cell death. Physiochemical properties of red pepper powder were not changed for one month after treatment with DBD plasma. Thus, DBD plasma treatment for 15 min is recommended to control foodborne pathogens in red pepper powder without changing its physicochemical properties. • Dielectric barrier discharge (DBD) plasma can enhance the quality of red pepper powder. • Effect of DBD plasma was more effective on E. coli O157: H7 than on B. cereus. • Expression levels of virulence genes of E. coli O157: H7 and B. cereus were decreased. • The morphological change of pathogens and red pepper powder were examined. [ABSTRACT FROM AUTHOR]

Over the past two decades, opioid abuse has risen especially among women. In both sexes hippocampal neural circuits involved in associative memory formation and encoding of motivational incentives are critically important in the transition from initial drug use to drug abuse/dependence. Opioid circuits, particularly the mossy fiber pathway, are crucial for associative memory processes important for addiction. Our anatomical studies, especially those utilizing electron microscopic immunocytochemistry, have provided unique insight into sex differences in the distribution of opioid peptides and receptors in specific hippocampal circuits and how these distributions are altered following stress and oxycodone-associative learning processes. Here we review the hippocampal opioid system in rodents with respect to ovarian hormones effects and baseline sex differences then sex differences following acute and chronic stress. Next, we review sex differences in the hippocampal opioid system in unstressed and chronically stressed rats following oxycodone conditioned place preference. We show that opioid peptides and receptors are distributed within hippocampal circuits in females with elevated estrogen states in a manner that would enhance sensitivity to endogenous and exogenous opioids. Moreover, chronic stress primes the opioid system in females in a manner that would promote opioid-associative learning processes. In contrast, chronic stress has limited effects on the opioid system in males and reduces its capacity to support opioid-mediated learning processes. Interestingly, acute stress appears to prime males for opioid associative learning. On a broader scale the findings highlighted in this review have important implications in understanding sex differences in opioid drug use and abuse. • High estrogens in females increase hippocampal opioid system to promote excitation. • Opioid expression and distribution favors enhanced plasticity in females. • Acute stress enhances opioid-mediated plasticity in males but not females. • Chronic stress reduces opioid-mediated plasticity in males but primed in females. • Sex-dependent redistribution during opioid place conditioning facilitates learning. [ABSTRACT FROM AUTHOR]

Abstract: Understanding the changes in host gene expression that occur with bacterial infection will help to elucidate the basis of molecular genetic control of disease resistance. The effect of infecting chicks with Salmonella enterica serovar Enteritidis on the RNA expression level of Toll-like receptor (TLR) genes, and the correlation between TLR RNA expression level and bacterial burden in the cecum and spleen of young birds was studied. Chicks from two advanced intercross lines were either infected or mock infected with S. enteritidis at 1 day of age. The RNA expression levels of TLR2, TLR4 and TLR5 genes were assessed by quantitative reverse transcriptase-PCR (qRT-PCR) in cecum and spleen tissues harvested at one week post-infection. Infected chicks had significant upregulation of TLR2 RNA expression in spleen, TLR4 RNA expression in both cecum and spleen, and downregulation of TLR5 RNA expression in cecum. Bacterial burden of S. enteritidis in infected birds was not correlated with TLR RNA expression level. Infecting chicks with S. enteritidis caused an increase in TLR2, TLR4 and TLR5 RNA expression level in spleen in males but not in females. The effect of sex on response to S. enteritidis infection suggests a role for TLR signaling pathways in sex-based modulation of immune response to pathogens. High correlation between TLR2 and TLR4 mRNA expression level in cecum of S. enteritidis infected birds suggests coordinated regulation or simultaneous stimulation of these genes by S. enteritidis. In conclusion, this study clearly showed that young chicks respond to S. enteritidis infection by upregulating TLR2, TLR4 RNA expression. The downregulation of TLR5 RNA expression was observed in cecum by S. enteritidis infection, which might be beneficial to protect host cells from overstimulation by bacterial flagellin. [Copyright &y& Elsevier]

Abstract: The endoribonuclease L (RNase L) is the effector of the 2-5A system, a major enzymatic pathway involved in the molecular mechanism of interferons (IFNs). RNase L is a very unusual nuclease with a complex mechanism of regulation. It is a latent enzyme, expressed in nearly every mammalian cell type. Its activation requires its binding to a small oligonucleotide, 2-5A. 2-5A is a series of unique 5′-triphosphorylated oligoadenylates with 2′-5′ phosphodiester bonds. By regulating viral and cellular RNA expression, RNase L plays an important role in the antiviral and antiproliferative activities of IFN and contributes to innate immunity and cell metabolism. The 2-5A/RNase L pathway is implicated in mediating apoptosis in response to viral infections and to several types of external stimuli. Several recent studies have suggested that RNase L could have a role in cancer biology and evidence of a tumor suppressor function of RNase L has emerged from studies on the genetics of hereditary prostate cancer. [Copyright &y& Elsevier]

Abstract: Paraquat (PQ)-induced pulmonary toxicity is characterized by initial development of pulmonary edema, infiltration of inflammatory cells, and damage to the alveolar epithelium, which may progress to severe fibrosis. However, the exact role of PQ in the progression of the pathogenesis has not been clearly established. To understand the mechanism of PQ in pulmonary toxicity, we developed an animal model of PQ-induced lung injury by intranasal instillation of PQ solution using C57Black/6J mice. Twenty microliters of PQ solution (0.01, 0.01, and 0.04mg/mouse) was applied through the nares, and the same amount of vehicle was applied in control mice. The pathological progression of lung pathology in our mouse model was very similar to that of patients suffering from PQ poisoning. The lungs of some animals exposed to PQ showed acute fulmination, resulting in death from 5 days post-exposure, but others showed a more protracted injury, resulting in typical pulmonary fibrosis at 3 weeks. Using this PQ-poisoned mouse model, we examined the gene expression at the initial destructive phase (within 5 days) that fibrosis has not completely developed. We prepared RNAs after 6h, 24h, and 5 days and examined the changes of the expression levels for 45 selected genes. The genes showing >2-fold increase at 6h or a time-dependent decrease during this experimental period may be the early markers for the destructive phase. These genes are Mt1, Mt2, Hmox1, Gcl, GR, IL-6, IL-13, Txn1, Fas, FasL, Lpin2, Mmp1a, Mmp12, Sfp-B, Sfp-D, CAT, EC-SOD, GST, and Pltp. On the other hand, the genes involved in the development of fibrosis, such as procollagen, Fn1, Eln, SMA, and Mmp9, Timp1 were significantly increased on day 5, not at 6h nor at 24h, after PQ treatment (the late marker). The genes showing a significant increase (Mmp3 and Mmp8) or decrease (VEGFA) at 24h and 5 days and not at 6h may be also the late markers. These changes in gene expression, which are equalled to functional activities of proteins, will be the targets for future studies focused on the development on PQ-induced pulmonary damage. [Copyright &y& Elsevier]

Abstract: Endogenous retroviral sequences (ERVs) are dynamic genomic components with profound influences on gene expression and genomic structure. Their extent of expression is not well known. Several broadly targeted real-time reverse transcription PCR (QPCRs) systems for surveillance of RNA expression of the major groups of human gammaretroviral ERVs were constructed. The highly conserved reverse transcriptase (RT) and integrase (IN) domains of the pol gene were used as targets for the PCRs, which were both probe-based (TaqMan®) and probe-less (SYBR® Green). Different levels of primer and probe degeneracy, with or without inosine, were tested. Several of the PCRs had sensitivities of a few HERV nucleic acid copies per PCR reaction. Specificities were approximately as expected from the fit of primers and probes. Gammaretroviral HERV RNA expression was studied in different human tissues. Each HERV group had a specific pattern of expression. HERV-E was highly expressed in testis, HERV-I/T in brain and testis, HERV-H in brain and testis, while HERV-W was highly expressed in placenta. Gammaretroviral RNA was not detected in plasma from 50 blood donors in saliva from 20 persons. In conclusion, a set of tools for investigation of gammaretroviral HERV RNA expression was created. [Copyright &y& Elsevier]

The protein kinase C (PKC) family of serine/threonine kinases plays a role in a variety of physiological and pathophysiological processes in the brain including development, synaptic plasticity, epilepsy, ischemia, and neuronal cell death. The subgroup of atypical PKCs (aPKCs) comprises of three members, PKCι/λ, PKCζ, and PKMζ, with high amino acid homology. We used specific RNA probes and in situ hybridization to determine the expression patterns of all the three isoforms in the adult mouse brain. PKCι and PKMζ were found to be broadly expressed in most of the cortex, the limbic system, and the thalamus. In contrast, PKCζ transcription was restricted to distinct forebrain areas and the cerebellum. Here we present a first comprehensive overview of isotype-specific aPKC distribution in the central nervous system, thereby providing a solid ground for further studies on the functional implications of the different aPKCs in the neuronal system. [Copyright &y& Elsevier]

Comparative gene-expression profiling is an important tool in understanding molecular signatures of complex diseases as well as the responses of cells and tissues to external factors. With increasing microarray data, disease-specific molecular patterns are emerging but the acquisition of these data is expensive, difficult to customize and not well standardized. Once genome-wide scans identify differentially expressed genes in a given disease, cheaper, more easily customized methods will be needed for evaluating the expression of these genes in large population samples. Here we describe a novel multiplex microsphere bead assay (MBA) to compare gene expression levels. To test this assay we evaluated the expression levels of four transcripts (BRCA1, MGB1, DLG1 and ACT1) in normal and cancerous mammary tissue. The results were consistent with those generated by quantitative real-time PCR. [Copyright &y& Elsevier]

Two chitin synthase (CHS) genes of the red flour beetle, Tribolium castaneum, were sequenced and their transcription patterns during development examined. By screening a BAC library of genomic DNA from T. castaneum (Tc) with a DNA probe encoding the catalytic domain of a putative Tribolium CHS, several clones that contained CHS genes were identified. Two distinct PCR products were amplified from these BAC clones and confirmed to be highly similar to CHS genes from other insects, nematodes and fungi. The DNA sequences of these genes, TcCHS1 and TcCHS2, were determined by amplification of overlapping PCR fragments from two of the BAC DNAs and mapped to different linkage groups. Each ORF was identified and full-length cDNAs were also amplified, cloned and sequenced. TcCHS1 and TcCHS2 encode transmembrane proteins of 1558 and 1464 amino acids, respectively. The TcCHS1 gene was found to use alternate exons, each encoding 59 amino acids, a feature not found in the TcCHS2 gene. During development, Tribolium expressed TcCHS1 predominantly in the embryonic and pupal stages, whereas TcCHS2 was prevalent in the late larval and adult stages. The alternate exon 8a of TcCHS1 was utilized over a much broader range of development than exon 8b. We propose that the two isoforms of the TcCHS1 enzyme are used predominantly for the formation of chitin in embryonic and pupal cuticles, whereas TcCHS2 is utilized primarily for the synthesis of peritrophic membrane-associated chitin in the midgut. [Copyright &y& Elsevier]

Three proteins, MsCP20, MsCP27 and MsCP36, that are secreted in greatest quantity into the pharate pupal cuticle of Manduca sexta () were purified and their amino acid sequences determined by mass spectrometry and Edman degradation. Although these proteins become sclerotized and insoluble in the pupal exoskeleton, their sequences contain features characteristic for proteins occurring in less sclerotized pliable cuticles, such as arthrodial membranes and soft larval cuticles. These proteins carry a secondary modification attached to a threonine residue, presumably an O-linked sugar moiety. cDNA clones of the genes for MsCP20, MsCP27 and MsCP36 were constructed from pharate pupal integument RNA. Close agreement was found between the amino acid sequences determined by Edman degradation and sequences deduced from the cDNA clones. The molecular masses determined by protein sequencing for MsCP20, MsCP27, and MsCP36 were 17,713, 17,448, and 29,582 Da, respectively, in close agreement with the masses deduced from the corresponding cDNA clones (17,711, 17,410, and 29,638 Da). Temporal expression analysis indicates that MsCP20 and MsCP36 transcripts are present at low levels early in the fifth larval stadium, followed by a large increase in abundance prior to pupal ecdysis. MsCP27 was not detected during development of the fifth larval instar, but its transcript, like those of MsCP20 and MsCP36, increased to a peak level just before pupal ecdysis. Only the MsCP36 transcript was detected in adults. These results support the hypothesis that these proteins are synthesized by the epidermis and are subsequently deposited into the cuticle during the larval–pupal transformation of M. sexta where they become sclerotized in the formation of pupal exocuticle. [Copyright &y& Elsevier]

Glycosyltransferases are enzymes that synthesize oligosaccharides, polysaccharides and glycoconjugates. One type of glycosyltransferase is chitin synthase, a very important enzyme in biology, which is utilized by insects, fungi, and other invertebrates to produce chitin, a polysaccharide of β-1,4-linked N-acetylglucosamine. Chitin is an important component of the insect’s exoskeletal cuticle and gut lining. To identify and characterize a chitin synthase gene of the tobacco hornworm, Manduca sexta, degenerate primers were designed from two highly conserved regions in fungal and nematode chitin synthase protein sequences and then used to amplify a similar region from Manduca cDNA. A full-length cDNA of 5152 nucleotides was assembled for the putative Manduca chitin synthase gene, MsCHS1, and sequencing of genomic DNA verified the contiguity of the sequence. The MsCHS1 cDNA has an ORF of 4692 nucleotides that encodes a transmembrane protein of 1564 amino acid residues with a mass of approximately 179 kDa (GenBank no. AY062175). It is most similar, over its entire length of protein sequence, to putative chitin synthases from other insects and nematodes, with 68% identity to enzymes from both the blow fly, Lucilia cuprina, and the fruit fly, Drosophila melanogaster. The similarity with fungal chitin synthases is restricted to the putative catalytic domain, and the MsCHS1 protein has, at equivalent positions, several amino acids that are essential for activity as revealed by mutagenesis of the fungal enzymes. A 5.3-kb transcript of MsCHS1 was identified by northern blot hybridization of RNA from larval epidermis, suggesting that the enzyme functions to make chitin deposited in the cuticle. Further examination by RT-PCR showed that MsCHS1 expression is regulated in the epidermis, with the amount of transcript increasing during phases of cuticle deposition. [Copyright &y& Elsevier]

• atp6 is an active region of mitochondrial genome recombination. • atp6 is a critical factor in regulating the cytoplasmic male sterility in kenaf. • A signal transit peptide was specific identified in atp6 of kenaf. • Insufficient energy supply was the main cause of kenaf cytoplasmic male sterility. Plant cytoplasmic male sterility (CMS) is a maternal genetic trait that results from incompatibilities between the organellar and nuclear genomes which causes pollen abortion. Cytoplasmic male sterility plays an important role in hybrid seed production and heterosis utilization. In kenaf, cytoplasmic male sterility has been exploited in hybrid seed production for several years, but its molecular mechanism has not yet been disclosed. In the present work, the mitochondrial genome of the isonuclear cytoplasmic male sterile line UG93A compared with maintainer line UG93B was described and a total of 398 single nucleotide polymorphisms (SNPs) and 230 insertions/deletions (InDels) were identified in the UG93A mitochondrial DNA. A total of 26 single nucleotide polymorphisms and insertions/deletions were identified in the coding region of atp6 , indicating active mitochondrial genome rearrangement. An expression cassette containing atp6 of UG93A without a fused mitochondrial transit peptide induced male sterility in tobacco and Arabidopsis. This indicated abnormal pollen grains and lack of pollen activity. Conclusively, coding sequence of atp6 , which contains single nucleotide polymorphism and indel variations may play a crucial role in regulating male sterility in kenaf. These data suggested that atp6 of UG93A may be a candidate gene for kenaf cytoplasmic male sterility and provided basic information for further study of the molecular mechanisms of cytoplasmic male sterility in kenaf. [ABSTRACT FROM AUTHOR]

• In mammals, exercise affects food intake. • Goldfish submitted to exercise had a lower post-exercise food intake compared to control fish. • Exercise increased hypothalamic expression levels of CART, IL6 and BDNF, but not orexin, irisin, CRF, leptin and NPY. • Exercise increased intestine CCK, PYY and GLP-1, and muscle irisin and IL-6 expression levels. • Exercise had no effects on expression levels of hepatic leptin and metabolic enzymes. In mammals, growing evidence indicates that exercise affects food intake, metabolism and the expression and blood levels of appetite regulators. In this study, we examined the effects of short-term (30 min, at low and high water flow) exercise on food intake, glucose levels and the expressions of appetite regulators in goldfish hypothalamus (irisin, orexin, CART, leptin), intestine (CCK, PYY, proglucagon/GLP-1), muscle (irisin) and liver (leptin), of brain-derived neurotrophic factor (BDNF) in brain, interleukin-6 (IL6) in muscle and hypothalamus, and major metabolic enzymes, the glycolytic enzyme glucokinase (GCK) and its regulatory protein (GCKR) in liver, the lipolytic enzyme lipoprotein lipase in intestine and muscle, and trypsin in intestine. Fish submitted to high flow exercise had a lower post-exercise food intake compared to control fish but no differences were seen in glucose levels between groups. Exercise induced an increase in hypothalamic expression levels of CART, IL6 and BDNF, but not orexin, irisin, CRF, leptin and NPY. High flow exercise induced an increase in intestine CCK, PYY and GLP-1, and muscle irisin and IL-6 expression levels. Exercise had no effects on expression levels of hepatic leptin or any of the metabolic enzymes examined. Our results suggest that, in goldfish, short-term exercise might decrease feeding in part by affecting the expressions of myokines and peripheral, but not central appetite regulators or metabolic enzyme/hormones. [ABSTRACT FROM AUTHOR]

The purpose of this research was to use RNA as a subject of legal medicine. Tissue samples were obtained from postmortem human brains collected from cadavers, whose reasons of death were exactly determined. Total RNA was isolated by the standard method and used for reverse transcription-polymerase chain reaction analysis (RT-PCR). The subject of our special concern was NOS and its expression in various brain tissues during some pathological processes. In this study, we showed that postmortem brain tissue can yield good quality mRNA for the successful application of traditional molecular biology methods from postmortem brain tissue even with prolonged postmortem intervals (up to 7 days). The relationship between a level of inducible NOS expression and intensity of pathomorphologic development in organs and tissues of people who died from different reasons was detected. [Copyright &y& Elsevier]