Validation of qPCR reference genes in lymphocytes from patients with amyotrophic lateral sclerosis

Usarek, Ewa; Barańczyk‐Kuźma, Anna; Kaźmierczak, Beata; Gajewska, Beata; Kuźma‐Kozakiewicz, Magdalena

doi:10.1371/journal.pone.0174317

Cited by 17 publications

(11 citation statements)

References 44 publications

(37 reference statements)

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“…We found that all analyzed RGs could be considered stable according to stability values calculated via the distinct algorithms, although there were some discrepancies according to the patients’ characteristics. These different results depending on the disease are in accordance with the work of Usarek et al, who found that the best RG for healthy PBMCs was B2M , whereas in PBMCs from patients with amyotrophic lateral sclerosis, it was HPRT1 [ 27 ]. Additionally, Wang et al found lower basal expression levels of PPIA in asthmatics than in healthy controls [ 15 ].…”

Section: Discussionsupporting

confidence: 88%

Reference Gene Validation for RT–qPCR in PBMCs from Asthmatic Patients with or without Obesity

Bantulà

Arismendi

Picado

et al. 2022

MPs

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Obesity is known to impair the efficacy of glucocorticoid medications for asthma control. Glucocorticoid-induced gene expression studies may be useful to discriminate those obese asthmatic patients who present a poor response to glucocorticoids. The expression of genes of interest is normalized with respect to reference genes (RGs). Ideally, RGs have a stable expression in different samples and are not affected by experimental conditions. The objective of this work was to analyze suitable RGs to study the role of glucocorticoid-induced genes in obese asthmatic patients in further research. The gene expression of eight potential RGs (GUSB, B2M, POLR2A, PPIA, ACTB, GAPDH, HPRT1, and TBP) was assessed with reverse transcription–quantitative polymerase chain reaction in peripheral blood mononuclear cells (PBMCs) from asthmatic, obese asthmatic, and healthy individuals. Their stability was analyzed using four different algorithms—BestKeeper, ΔCt, geNorm, and NormFinder. geNorm analysis recommended the use of a minimum of three genes for normalization. Moreover, intergroup variation due to the treatment was calculated by NormFinder, which found that B2M was the gene that was least affected by different treatments. Comprehensive rankings indicated GUSB and HPRT1 as the best RGs for qPCR in PBMCs from healthy and asthmatic subjects, while B2M and PPIA were the best for obese asthmatic subjects. Finally, our results demonstrated that B2M and HPRT1 were the most stable RGs among all groups, whereas ACTB, TBP, and GAPDH were the worst shared ones.

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Section: Discussionsupporting

confidence: 88%

Reference Gene Validation for RT–qPCR in PBMCs from Asthmatic Patients with or without Obesity

Bantulà

Arismendi

Picado

et al. 2022

MPs

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“…Upregulation of CTSH has also been reported in the spinal cord of SOD1 G93A mice but has not been studied thoroughly ( 59 ). Similarly, upregulated B2M has been reported in the peripheral blood of ALS patients ( 60 , 61 ). ITGB2, C3, C1QA, C1QB, and C1QC were observed as central genes in PPI analysis, and each of them performed an important role in synaptic pruning.…”

Section: Discussionsupporting

confidence: 56%

Identification of Molecular Correlations Between DHRS4 and Progressive Neurodegeneration in Amyotrophic Lateral Sclerosis By Gene Co-Expression Network Analysis

Zhu²,

Wei³

et al. 2022

Front. Immunol.

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Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease, and its candidate biomarkers have not yet been fully elucidated in previous studies. Therefore, with the present study, we aim to define and verify effective biomarkers of ALS by bioinformatics. Here, we employed differentially expressed gene (DEG) analysis, weighted gene co-expression network analysis (WGCNA), enrichment analysis, immune infiltration analysis, and protein–protein interaction (PPI) to identify biomarkers of ALS. To validate the biomarkers, we isolated the lumbar spinal cord from mice and characterized them using Western blotting and immunofluorescence. The results showed that Dhrs4 expression in the spinal cord was upregulated with the progression of SOD1G93A mice, and the upregulation of DHRS4 and its synergistic DHRS3 might be primarily associated with the activation of the complement cascade in the immune system (C1QA, C1QB, C1QC, C3, and ITGB2), which might be a novel mechanism that induces spinal neurodegeneration in ALS. We propose that DHRS4 and its synergistic DHRS3 are promising molecular markers for detecting ALS progression.

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“…Properdin, as well as Factor D, are the components of the complement system. They act as an activator of the complement system by stabilizing the alternative pathway convertase [ 46 , 47 ]. The alternative pathway of complement activation is associated with many autoimmune diseases, such as arthritis and age-related macular degeneration [ 48 ].…”

Section: Discussionmentioning

confidence: 99%

The Potential Role of Proinflammatory Cytokines and Complement Components in the Development of Drug-Induced Neuropathy in Patients with Multiple Myeloma

Łuczkowska

Rutka

Rogińska

et al. 2021

JCM

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The launch of novel chemotherapeutic agents—in particular, proteasome inhibitors and immunomodulatory drugs—dramatically changed multiple myeloma (MM) therapy, improving the response rate and prolonging progression-free survival. However, none of the anti-MM drugs are deprived of side effects. Peripheral neuropathy (PN) seems to be one of the most pressing problems. Despite extensive research in this area, the pathogenesis of drug-induced peripheral neuropathy (DiPN) has not yet been fully elucidated. In the present study, we aimed to assess the potential relationship between proinflammatory factors and the development of PN in MM patients with particular emphasis on the application of VTD (bortezomib, thalidomide, dexamethasone) regimen. Our analysis identified increased concentrations of CCL2, IL-1β, and IFN-γ in plasma of MM patients during treatment, both with and without symptoms of PN, compared with untreated neuropathy-free MM patients. At the same time, the plasma concentration of IL-1β in patients with neuropathy was significantly increased compared with patients without PN before and during treatment. Moreover, the results were enhanced at the transcript level by performing global mRNA expression analysis using microarray technology. The most significant changes were observed in the expression of genes responsible for regulating immunological and apoptotic processes. An in-depth understanding of the mechanisms responsible for the development of DiPN might in the future reduce the incidence of PN and accelerate diagnosis, allowing the choice of neuropathy-free treatment strategies for MM.

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Validation of qPCR reference genes in lymphocytes from patients with amyotrophic lateral sclerosis

Cited by 17 publications

References 44 publications

Reference Gene Validation for RT–qPCR in PBMCs from Asthmatic Patients with or without Obesity

Reference Gene Validation for RT–qPCR in PBMCs from Asthmatic Patients with or without Obesity

Identification of Molecular Correlations Between DHRS4 and Progressive Neurodegeneration in Amyotrophic Lateral Sclerosis By Gene Co-Expression Network Analysis

The Potential Role of Proinflammatory Cytokines and Complement Components in the Development of Drug-Induced Neuropathy in Patients with Multiple Myeloma

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