scholarly journals LC-MS/MS analysis of lesional and normally looking psoriatic skin reveals significant changes in protein metabolism and RNA processing

PLoS ONE ◽  
2021 ◽  
Vol 16 (5) ◽  
pp. e0240956
Author(s):  
V. V. Sobolev ◽  
A. V. Mezentsev ◽  
R. H. Ziganshin ◽  
A. G. Soboleva ◽  
M. Denieva ◽  
...  
Keyword(s):  
Healthy Volunteers ◽  
Rna Processing ◽  
Protein Identification ◽  
Expression Patterns ◽  
Differentially Expressed ◽  
Psoriatic Skin ◽  
Differentially Expressed Proteins ◽  
Kinin System ◽  
Lesional Skin ◽  
Kallikrein Kinin System

Background Plaque psoriasis is a chronic autoimmune disorder characterized by the development of red scaly plaques. To date psoriasis lesional skin transcriptome has been extensively studied, whereas only few proteomic studies of psoriatic skin are available. Aim The aim of this study was to compare protein expression patterns of lesional and normally looking skin of psoriasis patients with skin of the healthy volunteers, reveal differentially expressed proteins and identify changes in cell metabolism caused by the disease. Methods Skin samples of normally looking and lesional skin donated by psoriasis patients (n = 5) and samples of healthy skin donated by volunteers (n = 5) were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). After protein identification and data processing, the set of differentially expressed proteins was subjected to protein ontology analysis to characterize changes in biological processes, cell components and molecular functions in the patients’ skin compared to skin of the healthy volunteers. The expression of selected differentially expressed proteins was validated by ELISA and immunohistochemistry. Results The performed analysis identified 405 and 59 differentially expressed proteins in lesional and normally looking psoriatic skin compared to healthy control. In normally looking skin of the patients, we discovered decreased expression of KNG1, APOE, HRG, THBS1 and PLG. Presumably, these changes were needed to protect the epidermis from spontaneous activation of kallikrein-kinin system and delay the following development of inflammatory response. In lesional skin, we identified several large groups of proteins with coordinated expression. Mainly, these proteins were involved in different aspects of protein and RNA metabolism, namely ATP synthesis and consumption; intracellular trafficking of membrane-bound vesicles, pre-RNA processing, translation, chaperoning and degradation in proteasomes/immunoproteasomes. Conclusion Our findings explain the molecular basis of metabolic changes caused by disease in skin lesions, such as faster cell turnover and higher metabolic rate. They also indicate on downregulation of kallikrein-kinin system in normally looking skin of the patients that would be needed to delay exacerbation of the disease. Data are available via ProteomeXchange with identifier PXD021673.

scholarly journals LC-MS/MS analysis of lesional and normally looking psoriatic skin reveals significant changes in protein metabolism and RNA processing

2020 ◽  
Author(s):  
V.V. Sobolev ◽  
R.H. Ziganshin ◽  
A.V. Mezentsev ◽  
A.G. Soboleva ◽  
M. Denieva ◽  
...  
Keyword(s):  
Healthy Volunteers ◽  
Rna Processing ◽  
Protein Identification ◽  
Expression Patterns ◽  
Differentially Expressed ◽  
Psoriatic Skin ◽  
Differentially Expressed Proteins ◽  
Kinin System ◽  
Lesional Skin ◽  
Kallikrein Kinin System

AbstractBackgroundPlaque psoriasis is a chronic autoimmune disorder characterized by the development of red scaly plaques. To date psoriasis lesional skin transcriptome has been extensively studied, whereas only few proteomic studies of psoriatic skin are available.AimThe aim of this study was to compare protein expression patterns of lesional and normally looking skin of psoriasis patients with skin of the healthy volunteers, reveal differentially expressed proteins and identify changes in cell metabolism caused by the disease.MethodsSkin samples of normally looking and lesional skin donated by psoriasis patients (n = 5) and samples of healthy skin donated by volunteers (n = 5) were analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). After protein identification and data processing, the set of differentially expressed proteins was subjected to protein ontology analysis to characterize changes in biological processes, cell components and molecular functions in the patients’ skin compared to skin of the healthy volunteers.ResultsThe performed analysis identified 405 and 59 differentially expressed proteins in lesional and normally looking psoriatic skin compared to healthy control. We discovered decreased expression of KNG1, APOE, HRG, THBS1 and PLG in normally looking skin of the patients. Presumably, these changes were needed to protect the epidermis from spontaneous activation of kallikrein-kinin system and delay the following development of inflammatory response. In lesional skin, we identified several large groups of proteins with coordinated expression. Mainly, these proteins were involved in different aspects of protein and RNA metabolism, namely ATP synthesis and consumption; intracellular trafficking of membrane-bound vesicles, pre-RNA processing, translation, chaperoning and degradation in proteasomes/immunoproteasomes.ConclusionOur findings explain the molecular basis of metabolic changes caused by disease in skin lesions, such as faster cell turnover and higher metabolic rate. They also indicate on downregulation of kallikrein-kinin system in normally looking skin of the patients that would be needed to delay exacerbation of the disease. Data are available via ProteomeXchange with identifier PXD021673.

A Proteomic Approach to Discovery of Candidate Proteins Involved in the Transformation of Follicular Lymphoma.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 984-984
Author(s):  
Guang Fan ◽  
Yanping Zhong ◽  
Cristina Smith ◽  
James Huang ◽  
Rita Braziel
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Gel Electrophoresis ◽  
Western Blot Analysis ◽  
Protein Identification ◽  
Differentially Expressed ◽  
Signal Transduction Pathways ◽  
Differentially Expressed Proteins ◽  
2D Gel ◽  
2D Gel Analysis

Abstract Background: Follicular lymphoma (FL) undergoes transformation to a high grade diffuse large B-cell lymphoma (tr-DLBCL) in about 50% of patients. During transformation, a more virulent subclone of tumor cells emerges, leading to a rapidly progressive clinical course and resistance to therapy. The identification of proteins involved in transformation is critical for understanding the mechanism of transformation and developing molecularly targeted therapy. In this study, we compared protein expression between grade 1- FL (G1-FL) and tr-DLBCL using 2D-gel electrophoresis and Western blot analysis. Design: Frozen tissue and frozen cells were obtained from the Department of Pathology, Oregon Health and Science University tumor bank. The protein expression profiles of 3 G1-FL and 3 tr-DLBCL were compared using 2D-gel electrophoresis. Protein identification was done using a MALDI mass spectrometer. Frozen cells of an additional 11 non-paired GI-FL and 11 non-paired tr-DLBCL, and 2 pairs of G1-FL and tr-DLBCL specimens were used for Western blot confirmation of the initial 2D-gel findings. Results: 2D-gel analysis and MALDI protein identification revealed 14 differentially expressed proteins between G1-FL and tr-DLBCL (figure 1), all of which are known to play important roles in cellular energy/metabolic pathways, signal transduction pathways, and protein and nuclear synthesis. The two most differentially expressed proteins on 2D-gel analysis were superoxide dismutase (MnSOD2) and growth factor receptor bound protein 2 (Grb2). Western blot analysis of MnSOD2 and Grb2 confirmed their relative over- or under-expression in frozen cells from multiple additional clinical lymphoma samples, including 2 paired- and 22 non-paired G1-FL and tr-DLBCL. Both 2D-gel analysis and Western Blot showed a significantly higher level of expression of MnSOD2 and a lower expression of Grb2 expression in tr-DLBCL (figure 2). Summary: Using proteomic profiling, confirmed by Western blot analysis of clinical G1-FL and tr-DLBCL samples, we have confirmed 2 proteins (MnSOD2 and Grb2) that are expressed at significantly different levels in G1-FL and DLBCL. MnSOD2 is capable of protecting cells from reactive oxygen species and regulating signal transduction pathways to influence cell growth and apoptosis. Inhibition of MnSOD2 has been shown in studies of several cancer cell lines to render cancer cells more susceptible to apoptosis. Grb2 is a member of a critical signaling pathway leading to Ras activation in hematopoietic cells. Both proteins may play a critical role in FL transformation. These proteins have the potential to be therapeutic drug targets, diagnostic and/or prognostic markers, or biomarkers for monitoring therapeutic response. Summary of Differentially Expressed Spots Summary of Differentially Expressed Spots Figure Figure

Proteomic analysis of differentially expressed proteins in the roots of Columbia-0 and Landsberg erecta ecotypes of Arabidopsis thaliana in response to aluminum toxicity

2012 ◽  
Vol 92 (7) ◽  
pp. 1267-1282 ◽  
Author(s):  
T. Karuppanapandian ◽  
S-J. Rhee ◽  
E-J. Kim ◽  
B. K. Han ◽  
O. A. Hoekenga ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Stress Responses ◽  
Proteomic Analysis ◽  
Crop Production ◽  
Aluminum Toxicity ◽  
Expression Patterns ◽  
Differentially Expressed ◽  
Monodehydroascorbate Reductase ◽  
Differentially Expressed Proteins ◽  
Al Stress

Karuppanapandian, T., Rhee, S.-J., Kim, E.-J., Han, B. K., Hoekenga, O. A. and Lee, G. P. 2012. Proteomic analysis of differentially expressed proteins in the roots of Columbia-0 and Landsberg erecta ecotypes of Arabidopsis thaliana in response to aluminum-toxicity. Can. J. Plant Sci. 92: 1267–1282. Aluminum (Al) is phytotoxic when solubilized into Al3+ in acidic soils and represents a major constraint for crop production. The present study describes Al-stress responses in roots of Al-tolerant and Al-sensitive Arabidopsis ecotypes, Columbia-0 (Col-0) and Landsberg erecta (Ler), respectively. Comparative proteomic analysis was applied to plants grown in hydroponic solution culture under acidic pH (4.2) conditions. To investigate time-dependent responses, 6-d-old seedlings were treated with 30 µM AlCl3 for 24, 48, or 72 h; total proteins were prepared from roots and separated by two-dimensional gel electrophoresis (2-DE). From 2-DE analysis, were 600 proteins were inspected, 29 proteins were differentially responsive to Al-treatment. The 2-DE patterns were compared and differentially expressed proteins identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Analysis of protein expression patterns revealed that a set of proteins is functionally associated with tricarboxylic acid (TCA) cycle and glycolysis, reactive oxygen quenching and detoxification mechanism, and signal transduction pathways, etc., could play important roles in mediating plant response to Al in Arabidopsis ecotypes. Comparison of the changes in the protein profiles revealed that Al-stress increased Al-tolerance related proteins in Al-tolerant Col-0, but only generic stress responses occurred in Al-sensitive Ler. Specifically, Al up-regulated proteins such as alcohol dehydrogenase, monodehydroascorbate reductase, GTP-binding nuclear protein Ran-2, and leucine aminopeptidase in Col-0 but not in Ler.

scholarly journals Differential Expression of Estrogen-Responsive Genes in Women with Psoriasis

2021 ◽  
Vol 11 (9) ◽  
pp. 925
Author(s):  
Vladimir Sobolev ◽  
Anna Soboleva ◽  
Elena Denisova ◽  
Malika Denieva ◽  
Eugenia Dvoryankova ◽  
...  
Keyword(s):  
Healthy Volunteers ◽  
Differential Expression ◽  
Biological Effects ◽  
Differentially Expressed ◽  
Lesional Skin ◽  
Female Patients ◽  
Genes Encoding ◽  
Liquid Chromatography Tandem Mass ◽  
Severity Of The Disease ◽  
Skin Samples

In women, the flow of psoriasis is influenced by each phase of a woman’s life cycle. According to previous findings, significant changes in the levels of sex hormones affect the severity of the disease. Aim: The aim of this study was to identify the estrogen-responsive genes that could be responsible for the exacerbation of psoriasis in menopausal women. Methods: Skin samples of lesional skin donated by psoriasis patients (n = 5) were compared with skin samples of healthy volunteers (n = 5) using liquid chromatography–tandem mass spectrometry (LC–MS/MS). The set of differentially expressed proteins was subjected to protein ontology analysis to identify differentially expressed estrogen-responsive proteins. The expression of discovered proteins was validated by qPCR and ELISA on four groups of female participants. The first group included ten psoriasis patients without menopause; the second included eleven postmenopausal patients; the third included five healthy volunteers without menopause; and the fourth included six postmenopausal volunteers. Moreover, the participants’ blood samples were used to assess the levels of estradiol, progesterone, and testosterone. Results: We found that the levels of estradiol and progesterone were significantly lower and the levels of testosterone were significantly higher in the blood of patients compared to the control. The protein ontology analysis of LC–MS/MS data identified six proteins, namely HMOX1, KRT19, LDHA, HSPD1, MAPK1, and CA2, differentially expressed in the lesional skin of female patients compared to male patients. ELISA and qPCR experiments confirmed differential expression of the named proteins and their mRNA. The genes encoding the named proteins were differentially expressed in patients compared to volunteers. However, KRT19 and LDHA were not differentially expressed when we compared patients with and without menopause. All genes, except MAPK1, were differentially expressed in patients with menopause compared to the volunteers with menopause. HMOX1, KRT19, HSPD1, and LDHA were differentially expressed in patients without menopause compared to the volunteers without menopause. However, no significant changes were found when we compared healthy volunteers with and without menopause. Conclusion: Our experiments discovered a differential expression of six estrogen-controlled genes in the skin of female patients. Identification of these genes and assessment of the changes in their expression provide insight into the biological effects of estrogen in lesional skin. The results of proteomic analysis are available via ProteomeXchange with identifier PXD021673.

scholarly journals Expression Patterns of Clock Gene mRNAs and Clock Proteins in Human Psoriatic Skin Samples

2021 ◽  
Vol 23 (1) ◽  
pp. 121
Author(s):  
Viktória Németh ◽  
Szabina Horváth ◽  
Ágnes Kinyó ◽  
Rolland Gyulai ◽  
Zsuzsanna Lengyel
Keyword(s):  
Clock Gene ◽  
Clock Genes ◽  
Expression Patterns ◽  
Psoriatic Skin ◽  
Healthy Skin ◽  
Lesional Skin ◽  
Clock Proteins ◽  
Gene Transcripts ◽  
Skin Biopsies ◽  
Inflammatory Skin Disorder

Psoriasis is a systemic inflammatory skin disorder that can be associated with sleep disturbance and negatively influence the daily rhythm. The link between the pathomechanism of psoriasis and the circadian rhythm has been suggested by several previous studies. However, there are insufficient data on altered clock mechanisms in psoriasis to prove these theories. Therefore, we investigated the expression of the core clock genes in human psoriatic lesional and non-lesional skin and in human adult low calcium temperature (HaCaT) keratinocytes after stimulation with pro-inflammatory cytokines. Furthermore, we examined the clock proteins in skin biopsies from psoriatic patients by immunohistochemistry. We found that the clock gene transcripts were elevated in psoriatic lesions, especially in non-lesional psoriatic areas, except for rev-erbα, which was consistently downregulated in the psoriatic samples. In addition, the REV-ERBα protein showed a different epidermal distribution in non-lesional skin than in healthy skin. In cytokine-treated HaCaT cells, changes in the amplitude of the bmal1, cry1, rev-erbα and per1 mRNA oscillation were observed, especially after TNFα stimulation. In conclusion, in our study a perturbation of clock gene transcripts was observed in uninvolved and lesional psoriatic areas compared to healthy skin. These alterations may serve as therapeutic targets and facilitate the development of chronotherapeutic strategies in the future.

scholarly journals Targeted LC-MS/MS platform for the comprehensive determination of peptides in the kallikrein-kinin system

2021 ◽  
Author(s):  
Tanja Gangnus ◽  
Bjoern B. Burckhardt
Keyword(s):  
Healthy Volunteers ◽  
Clinical Symptoms ◽  
Matrix Effects ◽  
Accurate Determination ◽  
Kinin System ◽  
Carry Over ◽  
Kallikrein Kinin System ◽  
The Impact

AbstractThe kallikrein-kinin system (KKS) is involved in many physiological and pathophysiological processes and is assumed to be connected to the development of clinical symptoms of angioedema or COVID-19, among other diseases. However, despite its diverse role in the regulation of physiological and pathophysiological functions, knowledge about the KKS in vivo remains limited. The short half-lives of kinins, their low abundance and structural similarities and the artificial generation of the kinin bradykinin greatly hinder reliable and accurate determination of kinin levels in plasma. To address these issues, a sensitive LC-MS/MS platform for the comprehensive and simultaneous determination of the four active kinins bradykinin, kallidin, des-Arg(9)-bradykinin and des-Arg(10)-kallidin and their major metabolites bradykinin 2-9, bradykinin 1-7 and bradykinin 1-5 was developed. This platform was validated according to the bioanalytical guideline of the US Food and Drug Administration regarding linearity, accuracy, precision, sensitivity, carry-over, recovery, parallelism, matrix effects and stability in plasma of healthy volunteers. The validated platform encompassed a broad calibration curve range from 2.0–15.3 pg/mL (depending on the kinin) up to 1000 pg/mL, covering the expected concentrations in disease states. No source-dependent matrix effects were identified, and suitable stability of the analytes in plasma was observed. The applicability of the developed platform was proven by the determination of endogenous levels in healthy volunteers, whose plasma kinin levels were successfully detected in the low pg/mL range. The established platform facilitates the investigation of kinin-mediated diseases (e.g. angioedema, COVID-19) and enables the assessment of the impact of altered enzyme activities on the formation or degradation of kinins. Graphical abstract

scholarly journals Proteomic identification of glutamine synthetase as a differential marker for oligodendrogliomas and astrocytomas

2011 ◽  
Vol 115 (4) ◽  
pp. 789-795 ◽  
Author(s):  
Zhengping Zhuang ◽  
Meng Qi ◽  
Jie Li ◽  
Hiroaki Okamoto ◽  
David S. Xu ◽  
...  
Keyword(s):  
Glutamine Synthetase ◽  
Glial Cell ◽  
Cell Cultures ◽  
Gel Electrophoresis ◽  
Expression Patterns ◽  
Morphological Variability ◽  
Differentially Expressed ◽  
Differentially Expressed Proteins ◽  
Two Dimensional Gel Electrophoresis ◽  
Glial Cell Cultures

Object Astrocytomas and oligodendrogliomas are primary CNS tumors that remain a challenge to differentiate histologically because of their morphological variability and because there is a lack of reliable differential diagnostic markers. To identify proteins that are differentially expressed between astrocytomas and oligodendrogliomas, the authors analyzed the proteomic expression patterns and identified uniquely expressed proteins in these neoplasms. Methods Proteomes of astrocytomas and oligodendrogliomas were analyzed using 2D gel electrophoresis and subsequent computerized gel analysis to detect differentially expressed proteins. The proteins were identified using high-performance liquid chromatography accompanied by tandem mass spectrometry. To determine the role of the differentially expressed proteins in astrocytes, undifferentiated glial cell cultures were treated with dibutyryl–cyclic adenosine monophosphate (cAMP). Results Two-dimensional gel electrophoresis revealed that glutamine synthetase was differentially expressed in astrocytomas and oligodendrogliomas. Western blot and immunohistochemical analyses confirmed the increased expression of glutamine synthetase in astrocytomas compared with oligodendrogliomas. Whereas glutamine synthetase expression was demonstrated across all grades of astrocytomas (Grade II–IV [15 tumors]) and oligoastrocytomas (4 tumors), it was expressed in only 1 oligodendroglioma (6% [16 tumors]). Treatment of undifferentiated glial cell cultures with dibutyryl-cAMP resulted in astrocyte differentiation that was associated with increased levels of glial fibrillary acidic protein and glutamine synthetase. Conclusions These data indicate that glutamine synthetase expression can be used to distinguish astrocytic from oligodendroglial tumors and may play a role in the pathogenesis of astrocytomas.

scholarly journals High-throughput RNA sequencing from paired lesional- and non-lesional skin reveals major alterations in the psoriasis circRNAome

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Liviu-Ionut Moldovan ◽  
Thomas Birkballe Hansen ◽  
Morten Trillingsgaard Venø ◽  
Trine Line Hauge Okholm ◽  
Thomas Levin Andersen ◽  
...  
Keyword(s):  
Rna Sequencing ◽  
High Throughput ◽  
Mirna Expression ◽  
Binding Sites ◽  
Expression Patterns ◽  
Psoriatic Skin ◽  
Altered Expression ◽  
Lesional Skin ◽  
Inflammatory Skin ◽  
Laser Capture

Abstract Background Psoriasis is a chronic inflammatory skin disease characterized by hyperproliferation and abnormal differentiation of keratinocytes. It is one of the most prevalent chronic inflammatory skin conditions in adults worldwide, with a considerable negative impact on quality of life. Circular RNAs (circRNAs) are a recently identified type of non-coding RNA with diverse cellular functions related to their exceptional stability. In particular, some circRNAs can bind and regulate microRNAs (miRNAs), a group of RNAs that play a role in the pathogenesis of psoriasis. The aim of this study was to characterize the circRNAome in psoriasis and to assess potential correlations to miRNA expression patterns. Methods We used high-throughput RNA-sequencing (RNA-seq), NanoString nCounter technology and RNA chromogenic in situ hybridization (CISH) to profile the circRNA expression in paired lesional and non-lesional psoriatic skin from patients with psoriasis vulgaris. In addition, 799 miRNAs were profiled using NanoString nCounter technology and laser capture microdissection was used to study the dermis and epidermis separately. Results We found a substantial down-regulation of circRNA expression in lesional skin compared to non-lesional skin. We observed that this mainly applies to the epidermis by analyzing laser capture microdissected tissues. We also found that the majority of the circRNAs were downregulated independently of their corresponding linear host genes. The observed downregulation of circRNAs in psoriasis was neither due to altered expression levels of factors known to affect circRNA biogenesis, nor because lesional skin contained an increased number of inflammatory cells such as lymphocytes. Finally, we observed that the overall differences in available miRNA binding sites on the circRNAs between lesional and non-lesional skin did not correlate with differences in miRNA expression patterns. Conclusions We have performed the first genome-wide circRNA profiling of paired lesional and non-lesional skin from patients with psoriasis and revealed that circRNAs are much less abundant in the lesional samples. Whether this is a cause or a consequence of the disease remains to be revealed, however, we found no evidence that the loss of miRNA binding sites on the circRNAs could explain differences in miRNA expression between lesional and non-lesional skin.

scholarly journals Comparative Proteomics Reveals Cold Acclimation Machinery Through Enhanced Carbohydrate and Amino Acid Metabolism in Wucai (Brassica Campestris L.)

Plants ◽  
2019 ◽  
Vol 8 (11) ◽  
pp. 474
Author(s):  
Lingyun Yuan ◽  
Shilei Xie ◽  
Libing Nie ◽  
Yushan Zheng ◽  
Jie Wang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Cold Acclimation ◽  
Low Temperatures ◽  
Amino Acid Metabolism ◽  
Acid Metabolism ◽  
Expression Patterns ◽  
Differentially Expressed ◽  
Pcr Analysis ◽  
Differentially Expressed Proteins ◽  
Limited Information

Limited information is available on the cold acclimation of non-heading Chinese cabbage (NHCC) under low temperatures. In this study, the isobaric tags for relative and absolute quantification (iTRAQ) were used to illustrate the molecular machinery of cold acclimation. Compared to the control (Cont), altogether, 89 differentially expressed proteins (DEPs) were identified in wucai leaves responding to low temperatures (LT). Among these proteins, 35 proteins were up-regulated ((and 54 were down-regulated). These differentially expressed proteins were categorized as having roles in carbohydrate metabolism, photosynthesis and energy metabolism, oxidative defense, amino acid metabolism, metabolic progress, cold regulation, methylation progress, and signal transduction. The fructose, glucose, and sucrose were dramatically increased in response to cold acclimation. It was firstly reported that aspartate, serine, glutamate, proline, and threonine were significantly accumulated under low temperatures. Results of quantitative real-time PCR analysis of nine DEPs displayed that the transcriptional expression patterns of six genes were consistent with their protein expression abundance. Our results demonstrated that wucai acclimated to low temperatures through regulating the expression of several crucial proteins. Additionally, carbohydrate and amino acid conversion played indispensable and vital roles in improving cold assimilation in wucai.

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