scholarly journals Galactomannan and PCR in the Central Nervous System to Detect Invasive Mold Disease - A Retrospective Analysis in Immunocompromised Children

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Thomas Lehrnbecher ◽  
Peter Michael Rath ◽  
Andishe Attarbaschi ◽  
Gunnar Cario ◽  
Michaela Döring ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Infectious Complication ◽  
Pediatric Patients ◽  
Immunocompromised Patients ◽  
Chain Reaction ◽  
The Central Nervous System ◽  
Polymerase Chain ◽  
Pcr Assays ◽  
Work Up

Abstract Invasive mold disease (IMD) of the central nervous system (CNS) is a severe infectious complication in immunocompromised patients, but early microbiological diagnosis is difficult. As data on the value of biomarkers in the CNS are scarce, in particular in children, we retrospectively analyzed the performance of galactomannan (GM) and PCR assays in CNS samples of 15 children with proven and probable CNS IMD and of 32 immunocompromised children without fungal infection. Galactomannan in the cerebrospinal fluid (CSF) was assessed in nine of the 15 pediatric patients and was positive in five of them. Polymerase chain reaction (PCR) was performed in eight of the 15 patients and detected nucleic acids from molds in six patients. Galactomannan and PCR in CNS samples were the only positive microbiologic parameter in the CNS in three and two patients, respectively. In four patients, PCR specified the pathogen detected in microscopy. Galactomannan and PCR results remained negative in the CSF of all immunocompromised children without evidence for CNS IMD. Our data suggest that GM and PCR in CNS specimens are valuable additional tools in diagnosing CNS IMD and should be included in the work up of all pediatric patients with suspected mold disease of the CNS.

scholarly journals Transcriptional expression study in the central nervous system of rats: what gene should be used as internal control?

2014 ◽  
Vol 12 (3) ◽  
pp. 336-341 ◽  
Author(s):  
Ana Carolina de Moura ◽  
Virgínia Meneghini Lazzari ◽  
Grasiela Agnes ◽  
Silvana Almeida ◽  
Márcia Giovenardi ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Internal Control ◽  
Housekeeping Genes ◽  
Internal Control Gene ◽  
Chain Reaction ◽  
Beta Actin ◽  
The Central Nervous System ◽  
Polymerase Chain ◽  
The Brain

Objective A growing number of published articles report the expression of specific genes with different behavior patterns in rats. The levels of messenger ribonucleic acid transcripts are usually analyzed by reverse transcription followed by polymerase chain reaction and quantified after normalization with an internal control or reference gene (housekeeping gene). Nevertheless, housekeeping genes exhibit different expression in the central nervous system, depending on the physiological conditions and the area of the brain to be studied. The choice of a good internal control gene is essential for obtaining reliable results. This study evaluated the expression of three housekeeping genes (beta-actin, cyclophilin A, and ubiquitin C) in different areas of the central nervous system in rats (olfactory bulb, hippocampus, striatum, and prefrontal cortex). Methods Wistar rats (virgin females, n=6) during the diestrum period were used. Total ribonucleic acid was extracted from each region of the brain; the complementary deoxyribonucleic acid was synthesized by reverse transcription and amplified by real-time quantitative polymerase chain reaction using SYBR™ Green and primers specific for each one of the reference genes. The stability of the expression was determined using NormFinder. Results Beta-actin was the most stable gene in the hippocampus and striatum, while cyclophilin A and ubiquitin C showed greater stability in the prefrontal cortex and the olfactory bulb, respectively. Conclusion Based on our study, further studies of gene expression using rats as animal models should take into consideration these results when choosing a reliable internal control gene.

Expression of the integrin alpha 6 beta 4 in peripheral nerves: localization in Schwann and perineural cells and different variants of the beta 4 subunit

1994 ◽  
Vol 107 (2) ◽  
pp. 543-552 ◽  
Author(s):  
C.M. Niessen ◽  
O. Cremona ◽  
H. Daams ◽  
S. Ferraresi ◽  
A. Sonnenberg ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Polymerase Chain Reaction ◽  
Nervous System ◽  
Peripheral Nerves ◽  
Northern Blot Analysis ◽  
The Other ◽  
Chain Reaction ◽  
The Central Nervous System ◽  
Polymerase Chain ◽  
Integrin Alpha 6

Integrin alpha 6 beta 4 is expressed in human peripheral nerves, but not in the central nervous system. This integrin heterodimer has previously been found in perineural fibroblast-like cells and in Schwann cells (SCs), which both assemble a basement membrane but do not form hemidesmosomes. We show here that in SCs, which had formed a myelin sheath, alpha 6 beta 4 was enriched in the proximity of the nucleus, at Ranvier paranodal areas and at Schmitt-Lanterman clefts; alpha 6 beta 4 was also found at the grooved interface between small axons and non-myelinating SCs. Immunoprecipitation of human peripheral nerves, in combination with Western blotting showed that beta 4 is associated with the alpha 6A subunit. Northern blot analysis of human peripheral nerves showed a single beta 4 transcript of 6 kb. Using the reverse transcriptase polymerase chain reaction, we detected two mRNA species, one for the most common (−70, -53) form of beta 4 and the other encoding the (+53) variant of beta 4. Cultured SCs were devoid of alpha 6 beta 4 but expressed alpha 6 beta 1, indicating that SCs lose beta 4 expression when contact with neurons is lost. Thus, resting SCs in contact with axons express alpha 6A in combination with beta 4, irrespective of myelin formation. We suggest that alpha 6 beta 4 expressed in SCs plays a role in peripheral neurogenesis.

scholarly journals Quantitative RT-PCR Analysis of Uncoupling Protein Isoforms in Mouse Brain Cortex: Methodological Optimization and Comparison of Expression with Brown Adipose Tissue and Skeletal Muscle

2004 ◽  
Vol 24 (7) ◽  
pp. 780-788 ◽  
Author(s):  
Sylvain Lengacher ◽  
Pierre J. Magistretti ◽  
Luc Pellerin
Keyword(s):  
Skeletal Muscle ◽  
Central Nervous System ◽  
Polymerase Chain Reaction ◽  
Nervous System ◽  
Adipose Tissue ◽  
Brain Cortex ◽  
Chain Reaction ◽  
Brown Adipose ◽  
The Central Nervous System ◽  
Polymerase Chain

Uncoupling proteins (UCPs) present in the inner mitochondrial membrane are involved in uncoupling respiration from ATP synthesis. Five UCP isoforms have been identified but information about their presence and level of expression in the central nervous system remains incomplete. To determine the nature and proportion of UCP isoform mRNAs present in brain cortex, we developed and optimized a specific quantitative reverse-transcription polymerase chain reaction procedure. Optimal range of RNA concentrations to be used in the reverse-transcriptase reaction was determined. Primer design and concentration were optimized for each target gene while polymerase chain reaction efficiency was assessed for a range of reverse-transcriptase dilutions. Genomic contribution to the quantitative signal was evaluated for each isoform and minimized. Three reference genes were tested for normalization, and β-actin was found to be the most stable among tissues. Results indicate that brain cortex contains significant amounts of all UCP mRNAs, with UCP5 and UCP4 being the most abundant, as opposed to brown adipose tissue and skeletal muscle, which predominantly express UCP1 and UCP3, respectively. These data provide a first quantitative assessment of UCP mRNA expression in mouse brain, showing the presence of all five isoforms with distinct proportions, thus suggesting specific roles in the central nervous system.

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