Comparison of two FFPE preparation methods using label-free shotgun proteomics: Application to tissues of diverticulitis patients.
QUESADA CALVO, Florence ; Bertrand, Virginie ; Longuespée, Rémi et al
in Journal of proteomics (2014), 112C
Formalin-fixed paraffin-embedded (FFPE) specimens of patients are useful sources of materials for clinical research and have recently gained interest for use in the discovery of clinical proteomic ... [more ▼]
Formalin-fixed paraffin-embedded (FFPE) specimens of patients are useful sources of materials for clinical research and have recently gained interest for use in the discovery of clinical proteomic biomarkers. However, the critical step in this field is the ability to obtain an efficient and repeatable extraction using the limited quantities of material available for research in hospital biobanks. This work describes the evaluation of the peptide/protein extraction using FFPE sections treated by the following two methods before shotgun proteomic analysis: a commercial solution (FFPE-FASP) (filter aided sample preparation) and an antigen retrieval-derived protocol (On Slice AR). Their efficiencies and repeatabilities are compared using data-independent differential quantitative label-free analysis. FFPE-FASP was shown to be globally better both qualitatively and quantitatively than On Slice AR. FFPE-FASP was tested on several samples, and differential analysis was used to compare the tissues of diverticulitis patients (healthy and inflammatory tissues). In this differential proteomic analysis using retrospective clinical FFPE material, FFPE-FASP was reproducible and provided a high number of confident protein identifications, highlighting potential protein biomarkers. BIOLOGICAL SIGNIFICANCE: In clinical proteomics, FFPE is an important resource for retrospective analysis and for the discovery of biomarkers. The challenge for FFPE shotgun proteomic analysis is preparation by an efficient and reproducible protocol, which includes protein extraction and digestion. In this study, we analyzed two different methods and evaluated their repeatabilities and efficiencies. We illustrated the reproducibility of the most efficient method, FFPE-FASP, by a pilot study on diverticulitis tissue and on FFPE samples amount accessible in hospital biobanks. These data showed that FFPE is suitable for use in clinical proteomics, especially when the FFPE-FASP method is combined with label-free shotgun proteomics as described in the workflow presented in this work. [less ▲]Detailed reference viewed: 85 (28 ULg)
Vieillissement et cancer: coincidence ou relation etiologique?
; Demoulin, Stéphanie ; Petermans, Jean et al
in Revue medicale de Liege (2014), 69(5-6), 276-81
In Belgium and in other countries, the Cancer Registry data show an increased incidence of cancers related to age, the majority of tumors being diagnosed beyond 60 years. However, the mechanisms ... [more ▼]
In Belgium and in other countries, the Cancer Registry data show an increased incidence of cancers related to age, the majority of tumors being diagnosed beyond 60 years. However, the mechanisms responsible for this increase are not clear. Cancer could be chronologically associated with aging because of the long latency period between the exposition to carcinogenic agents and the appearance of clinical signs. Aging could also predispose directly to cancer by different mechanisms (impaired immune response, increased oxidative stress, shortening of telomeres, accumulation of senescent cells). In this review, we propose to describe different hypotheses which could explain the increased incidence of cancers in the elderly. [less ▲]Detailed reference viewed: 61 (0 ULg)
DeltaNp63 isoform-mediated beta-defensin family up-regulation is associated with (lymph)angiogenesis and poor prognosis in patients with squamous cell carcinoma.
Suarez-Carmona, Meggy ; Hubert, Pascale ; Gonzalez, Arnaud et al
in Oncotarget (2014), 5(7), 1856-1868
Beside a role in normal development/differentiation, high p63 immunoreactivity is also frequently observed in squamous cell carcinoma (SCC). Due to the complexity of the gene, the role of each p63 isotype ... [more ▼]
Beside a role in normal development/differentiation, high p63 immunoreactivity is also frequently observed in squamous cell carcinoma (SCC). Due to the complexity of the gene, the role of each p63 isotype in tumorigenesis is still confusing. Constitutively produced or induced in inflammatory conditions, human beta-defensins (HbetaDs) are cationic peptides involved in host defenses against bacteria, viruses and fungi. Here, we investigated both the role of p63 proteins in the regulation of HbetaDs and the implication of these antimicrobial peptides in tumor (lymph)angiogenesis. Thus, in contrast to TAp63 isotypes, we observed that DeltaNp63 proteins (alpha, beta, gamma) induce HbetaD1, 2 and 4 expression. Similar results were observed in cancer tissues and cell lines. We next demonstrated that DeltaNp63-overexpressing SCC are associated with both a poor prognosis and a high tumor vascularisation and lymphangiogenesis. Moreover, we showed that HbetaDs exert a chemotactic activity for (lymphatic) endothelial cells in a CCR6-dependent manner. The ability of HbetaDs to enhance (lymph)angiogenesis in vivo was also evaluated. We observed that HbetaDs increase the vessel number and induce a significant increase in relative vascular area compared to negative control. Taken together, the results of this study suggest that DeltaNp63-regulated HbetaD could promote tumor (lymph)angiogenesis in SCC microenvironment. [less ▲]Detailed reference viewed: 139 (13 ULg)
Accessibilome of human glioblastoma: collagen-VI-alpha-1 is a new target and a marker of poor outcome
Turtoi, Andrei ; Blomme, Arnaud ; BIANCHI, Elettra et al
in Journal of Proteome Research (2014), 13(12), 5660-5669
Functional targeted therapy has unfortunately failed to improve the outcome of glioblastoma patients. Success stories evidenced by the use of antibody-drug conjugates in other tumor types are encouraging ... [more ▼]
Functional targeted therapy has unfortunately failed to improve the outcome of glioblastoma patients. Success stories evidenced by the use of antibody-drug conjugates in other tumor types are encouraging, but targets specific to glioblastoma and accessible through the bloodstream remain scarce. In the current work, we have identified and characterized novel and accessible proteins using an innovative proteomic approach on six human glioblastomas; the corresponding data have been deposited in the PRIDE database identifier PXD001398. Among several clusters of uniquely expressed proteins, we highlight collagen-VI-alpha-1 (COL6A1) as a highly expressed tumor biomarker with low levels in most normal tissues. Immunohistochemical analysis of glioma samples from 61 patients demonstrated that COL6A1 is a significant and consistent feature of high-grade glioma. Deposits of COL6A1 were evidenced in the perivascular regions of the tumor-associated vasculature and in glioma cells found in pseudopalisade structures. Retrospective analysis of public gene-expression data sets from over 300 glioma patients demonstrated a significant correlation of poor patient outcome and high COL6A1 expression. In a proof-of-concept study, we use chicken chorioallantoic membrane in vivo model to show that COL6A1 is a reachable target for IV-injected antibodies. The present data warrant further development of human COL6A1 antibodies for assessing the quantitative biodistribution in the preclinical tumor models. [less ▲]Detailed reference viewed: 21 (3 ULg)
Le syndrome psoriasis avec ses comorbidités.
Franchimont, Claudine ; PIERARD, Gérald ; DELVENNE, Philippe et al
in Revue Médicale de Liège (2014), 69
The recognition of a complex disorder associating skin psoriasis and its comorbidities has considerably progressed over recent years. Beyond the skin lesions, psoriatic arthritis, the metabolic syndrome ... [more ▼]
The recognition of a complex disorder associating skin psoriasis and its comorbidities has considerably progressed over recent years. Beyond the skin lesions, psoriatic arthritis, the metabolic syndrome, type II diabetes, cardiovascular disease, inflammatory bowel diseases, and some cancers represent a group of chronic systemic disorders. The process is initiated and sustained by an immunological pathway involving Th1, Th17 and Th22 lymphocytes. The recognition of this complex disorder indicates that the clinical impact of psoriasis is not exclusively limited to the skin only. Keywords : Psoriasis - Cardiovascular disease [less ▲]Detailed reference viewed: 14 (0 ULg)
Altered alpha-defensin 5 expression in cervical squamocolumnar junction: implication in the formation of a viral/tumour-permissive microenvironment.
Hubert, Pascale ; ; RONCARATI, Patrick et al
in Journal of Pathology (The) (2014), 234(4), 464-77
Human papillomavirus (HPV) infection, particularly type 16, is causally associated with cancer of the uterine cervix, which mainly develops at the squamocolumnar (SC) junction. The progression of cervical ... [more ▼]
Human papillomavirus (HPV) infection, particularly type 16, is causally associated with cancer of the uterine cervix, which mainly develops at the squamocolumnar (SC) junction. The progression of cervical HPV infections into (pre)neoplastic lesions suggests that viral antigens are not adequately recognized by innate immunity or presented to the adaptive immune system. Members of the defensin family have recently been found to inhibit viral and bacterial pathogens, to stimulate the migration of immune cells and to play a role in anticancer responses. In the present study, we focused on the poorly characterized human alpha-defensin 5 (HD-5) and its possible role in these processes. We showed that HD-5 was able to prevent HPV virion entry into cervical keratinocytes and to influence adaptive immunity. Indeed, this peptide specifically induced the chemoattraction and proliferation of both activated T lymphocytes and immature dendritic cells in a CCR2/CCR6-dependent manner and stimulated the infiltration of these professional antigen-presenting cells in a (pre)neoplastic epithelium transplanted in vivo in immunodeficient mice. No chemotactic effect was observed with plasmacytoid dendritic cells, macrophages or natural killer cells. Proliferative and angiogenic effects of HD-5 were also assessed in vitro and in vivo. However there was a striking regional disparity in expression of HD-5, being prominent in ectocervical, vaginal and vulvar neoplasia, while absent, or nearly so, in the cervical SC junction. Taken together, these results suggest one possible explanation for why the SC junction is uniquely vulnerable to both high-risk HPV infection (via reduced HD-5 expression and viral entry) and progression of neoplasia (via altered cell-mediated immune responses and altered microenvironment). Copyright (c) 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [less ▲]Detailed reference viewed: 51 (14 ULg)
Dormancy of growth-stunted malignant melanoma: sustainable and smoldering patterns.
FRANCHIMONT, Claudine ; Hermanns-Lê, Trinh ; Delvenne, Philippe et al
in Oncology Reviews (2014), 8Detailed reference viewed: 35 (0 ULg)
Evaluation of CellSolutions BestPrep(R) Automated Thin-Layer Liquid-Based Cytology Papanicolaou Slide Preparation and BestCyte(R) Cell Sorter Imaging System.
; GOFFIN, Frédéric ; Kridelka, Frédéric et al
in Acta cytologica (2014)
Objective: A double-blind study was conducted to compare the performance of the new BestPrep(R) (CellSolutions) liquid-based thin-layer Papanicolaou (Pap) test with ThinPrep(R) (Hologic). Study Design ... [more ▼]
Objective: A double-blind study was conducted to compare the performance of the new BestPrep(R) (CellSolutions) liquid-based thin-layer Papanicolaou (Pap) test with ThinPrep(R) (Hologic). Study Design: Samples from the study patients (n = 105) were collected twice in the same encounter with the ThinPrep sample always taken first and the BestPrep sample collected second. Slides were prepared according to both manufacturers' protocols and evaluated using manual microscopic review and the BestCyte(R) cell sorter imaging system (CellSolutions). Diagnostic truth for each case was determined by independent manual review of both slides by multiple pathologists and histology when available. The presence of atypical squamous cells of undetermined significance was the threshold for positive for sensitivity and specificity calculations. Results: BestPrep and ThinPrep, by manual review, had sensitivities for high-grade squamous intraepithelial lesion (HSIL) cases of 100 and 95.6%, respectively. Using the BestCyte cell sorter, both had 100% sensitivity. For the same HSIL cases, the digene HC2 high-risk human papillomavirus DNA test had sensitivities of 100% (BestPrep) and 95.6% (ThinPrep). Specificities were 71.4% (BestPrep) and 54.8% (ThinPrep). Conclusions: BestPrep was equivalent to ThinPrep for manual review even though BestPrep was always the second sample collected. The BestCyte cell sorter provides a practical alternative to manual review for both BestPrep and ThinPrep slides. (c) 2014 S. Karger AG, Basel. [less ▲]Detailed reference viewed: 89 (7 ULg)
NF-kappaB-induced KIAA1199 promotes survival through EGFR signalling.
Shostak, Kateryna ; ; Hubert, Pascale et al
in Nature communications (2014), 5
Constitutive activation of EGFR- and NF-kappaB-dependent pathways is a hallmark of cancer, yet signalling proteins that connect both oncogenic cascades are poorly characterized. Here we define KIAA1199 as ... [more ▼]
Constitutive activation of EGFR- and NF-kappaB-dependent pathways is a hallmark of cancer, yet signalling proteins that connect both oncogenic cascades are poorly characterized. Here we define KIAA1199 as a BCL-3- and p65-dependent gene in transformed keratinocytes. KIAA1199 expression is enhanced on human papillomavirus (HPV) infection and is aberrantly expressed in clinical cases of cervical (pre)neoplastic lesions. Mechanistically, KIAA1199 binds Plexin A2 and protects from Semaphorin 3A-mediated cell death by promoting EGFR stability and signalling. Moreover, KIAA1199 is an EGFR-binding protein and KIAA1199 deficiency impairs EGF-dependent Src, MEK1 and ERK1/2 phosphorylations. Therefore, EGFR stability and signalling to downstream kinases requires KIAA1199. As such, KIAA1199 promotes EGF-mediated epithelial-mesenchymal transition (EMT). Taken together, our data define KIAA1199 as an oncogenic protein induced by HPV infection and constitutive NF-kappaB activity that transmits pro-survival and invasive signals through EGFR signalling. [less ▲]Detailed reference viewed: 41 (6 ULg)
Downscalling tissue proteomics: analyses of limited amounts of FFPE tissue samples. Application to cervix cancer biomarker markers.
Longuespée, Rémi ; ; Smargiasso, Nicolas et al
Poster (2014)Detailed reference viewed: 29 (9 ULg)
Cyanoacrylate skin surface strippings and the 3S-Bioket advent in tropical dermatology. A look from Liège.
PIERARD, Gérald ; Franchimont, Claudine ; PAQUET, Philippe et al
in Scientific World Journal (The) (2014), 2014
In the dermatopathology field, some simple available laboratory tests require minimum equipment for establishing a diagnosis. Among them, the cyanoacrylate skin surface stripping (CSSS), formerly named ... [more ▼]
In the dermatopathology field, some simple available laboratory tests require minimum equipment for establishing a diagnosis. Among them, the cyanoacrylate skin surface stripping (CSSS), formerly named skin surface biopsy or follicular biopsy, represents a convenient low cost procedure. It is a minimally invasive method collecting a continuous sheet of stratum corneum and horny follicular casts. In the vast majority of cases, it is painless and is unassociated with adverse events. CSSS can be performed in subjects of any age. The method has a number of applications in diagnostic dermatopathology and cosmetology, as well as in experimental dermatology settings. A series of derived analytic procedures include xerosis grading, comedometry, corneofungimetry, corneodynamics of stratum corneum renewal, corneomelametry, corneosurfametry, and corneoxenometry. [less ▲]Detailed reference viewed: 22 (8 ULg)
Tissue Proteomics for the Next Decade? Towards a Molecular Dimension in Histology
Longuespée, Rémi ; ; et al
in OMICS : A Journal of Integrative Biology (2014)Detailed reference viewed: 37 (8 ULg)
La dermatoporose, un "vintage" de l'atrophodermie et de la "peau transparente".
FRANCHIMONT, Claudine ; ; Hermanns-Lê, Trinh et al
in Revue Médicale de Liège (2014), 69
Le vieillissement de la peau se marque à terme par une atrophodermie, parfois sévère. La corticothérapie, qu’elle soit systémique ou topique, exerce un effet atrophiant qui copie l’effet du grand âge. Cet ... [more ▼]
Le vieillissement de la peau se marque à terme par une atrophodermie, parfois sévère. La corticothérapie, qu’elle soit systémique ou topique, exerce un effet atrophiant qui copie l’effet du grand âge. Cet état d’atrophodermie a été décrit comme la «peau transparente» il y a une quarantaine d’années, et il a reçu plus récemment la nouvelle dénomination de dermatoporose. Il est connu de longue date que cet état entraîne des conséquences cliniques parfois majeures qui conduisent à des lésions diverses sous forme de purpura de Bateman, de cicatrices stellaires, et de plaies faisant suite à des traumatismes mineurs. [less ▲]Detailed reference viewed: 46 (1 ULg)
Gynecologic and obstetric impact of the Ehlers-Danlos syndrome: clues from scrutinizing dermal ultrastructural alterations.
Hermanns-Lê, Trinh ; PIERARD, Gérald ; FRANCHIMONT, Claudine et al
in Gynecology (2014), 2(1),
Background: The Ehlers-Danlos syndrome (EDS) represents a heterogenous group of heritable connective tissue disorders chiefly characterized by joint laxity, increased skin distensibility and connective ... [more ▼]
Background: The Ehlers-Danlos syndrome (EDS) represents a heterogenous group of heritable connective tissue disorders chiefly characterized by joint laxity, increased skin distensibility and connective tissue fragility. Each EDS type corresponds to a multisystemic disorder with widespread implications. Physicians evoke the EDS diagnosis in presence of some typical cutaneous signs, such as atrophic scars and frequent hematomas. Methods: To revisit the transmission electron microscopy support from skin biopsies when screening EDS in gynecology-obstetrics. Results: EDS women commonly suffer from a series of gynecologic and obstetric disorders. Dermal ultrastructural changes are keynote features suggesting some EDS types. Conclusion: Gene mutations are mostly identified in the classic and vascular EDS types, but only in sporadic cases of the hypermobile EDS type which apparently represents the most frequent type. Dermal ultrastructural changes represent relevant diagnostic clues for the practitioner. Preconception counseling about possible complications is welcome. Management requires precise diagnosis of the EDS type. [less ▲]Detailed reference viewed: 48 (6 ULg)
Effect of N-acetylcysteine combined with infliximab on toxic epidermal necrolysis. A proof-of-concept study.
PAQUET, Philippe ; ; ROUSSEAU, Anne-Françoise et al
in Burns : Journal of the International Society for Burn Injuries (2014)Detailed reference viewed: 36 (4 ULg)
Le point de rosée en berne et la couche cornée hivernale.
PIERARD, Gérald ; FRANCHIMONT, Claudine ; Delvenne, Philippe
in Revue Médicale de Liège (2014), 69(2), 68-71
The frontier between the human body and the environment is largely represented by the stratum corneum. It is obvious that the skin aspect on some body sites appears dry and rough in some predisposed ... [more ▼]
The frontier between the human body and the environment is largely represented by the stratum corneum. It is obvious that the skin aspect on some body sites appears dry and rough in some predisposed individuals during winter season. The reductions in the outdoor temperature, ambient moisture and dew point are the main physical factors involved in this process. We present the aspects seen under ultraviolet light and scanning electron microscopy. Keywords : Stratum corneum - Dew point [less ▲]Detailed reference viewed: 18 (3 ULg)
Identification of a microRNA landscape targeting the PI3K/Akt signaling pathway in inflammation-induced colorectal carcinogenesis
JOSSE, Claire ; Bouznad, Nassim ; Geurts, Pierre et al
in American Journal of Physiology - Gastrointestinal and Liver Physiology (2014), 306
Inflammation can contribute to tumor formation; however, markers that predict progression are still lacking. In the present study, the well-established azoxymethane (AOM)/dextran sulfate sodium (DSS ... [more ▼]
Inflammation can contribute to tumor formation; however, markers that predict progression are still lacking. In the present study, the well-established azoxymethane (AOM)/dextran sulfate sodium (DSS)-induced mouse model of colitis-associated cancer was used to analyze microRNA (miRNA) modulation accompanying inflammation-induced tumor development and to determine whether inflammation-triggered miRNA alterations affect the expression of genes or pathways involved in cancer. A miRNA microarray experiment was performed to establish miRNA expression profiles in mouse colon at early and late time points during inflammation and/or tumor growth. Chronic inflammation and carcinogenesis were associated with distinct changes in miRNA expression. Nevertheless, prediction algorithms of miRNA-mRNA interactions and computational analyses based on ranked miRNA lists consistently identified putative target genes that play essential roles in tumor growth or that belong to key carcinogenesis-related signaling pathways. We identified PI3K/Akt and the insulin growth factor-1 (IGF-1) as major pathways being affected in the AOM/DSS model. DSS-induced chronic inflammation downregulates miR-133a and miR-143/145, which is reportedly associated with human colorectal cancer and PI3K/Akt activation. Accordingly, conditioned medium from inflammatory cells decreases the expression of these miRNA in colorectal adenocarcinoma Caco-2 cells. Overexpression of miR-223, one of the main miRNA showing strong upregulation during AOM/DSS tumor growth, inhibited Akt phosphorylation and IGF-1R expression in these cells. Cell sorting from mouse colons delineated distinct miRNA expression patterns in epithelial and myeloid cells during the periods preceding and spanning tumor growth. Hence, cell-type-specific miRNA dysregulation and subsequent PI3K/Akt activation may be involved in the transition from intestinal inflammation to cancer. [less ▲]Detailed reference viewed: 101 (18 ULg)