scholarly journals Keloid-Specific Gene Expression Profiling for Accurate Diagnostic and Therapeutic Applications

2021 ◽  
Author(s):  
Miae Choi ◽  
Jaemyun Lyu ◽  
Karthika Muthuramalingam ◽  
Eunmyong Lee ◽  
Yoojoung Oh ◽  
...  
Keyword(s):  
Normal Skin ◽  
Treatment Modalities ◽  
Specific Gene ◽  
Hypertrophic Scars ◽  
Transcriptome Profile ◽  
Normal Tissues ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed ◽  
Distinct Subtype

Abstract Scars are a heterogeneous disease including normotrophic scars, hypertrophic scars, and keloids. Of these lesions, keloids are a distinct subtype from any other type of scar. Clinically, it causes pain, itching, or tenderness, causing life discomfort and characteristically irreversible. Despite various treatment modalities, restoring keloids to normal tissues is difficult, and frequent recurrences have been reported. Therefore, it is essential to identify keloid-specific genes for accurate diagnosis and treatment of keloids. In an effort to find out keloid-specific genes, several studies compared keloids with scar-free normal skin, which leading general scar-related genes to be chosen rather than keloid-specific genes. To select for highly accurate keloid-specific genes and pathways, we compared the transcriptome profile of keloids with those of normotrophic scars and hypertrophic scars, which acquired from formalin-fixed paraffin-embedded human skin samples using high-throughput RNA-sequencing techniques. Differential expression analyses and over-representation analyses revealed that genes related to nervous system process were upregulated in keloids, whereas genes related with immune responses were downregulated in keloids. Additionally, the extracellular matrix related processes were highlighted in both hypertrophic scars and keloids. Finally, we highlight potential keloid-specific biomarkers and expression changes that can be employed for future therapeutics of keloids.

scholarly journals Specificity and sensitivity of immunohistochemical detection of factor VIII/von Willebrand factor antigen in formalin-fixed paraffin-embedded tissue.

1982 ◽  
Vol 30 (4) ◽  
pp. 371-377 ◽  
Author(s):  
R D McComb ◽  
T R Jones ◽  
S V Pizzo ◽  
D D Bigner
Keyword(s):  
Endothelial Cells ◽  
Von Willebrand Factor ◽  
Immunohistochemical Detection ◽  
Normal Tissues ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Von Willebrand ◽  
Willebrand Factor ◽  
Factor Antigen ◽  
Formalin Fixed

The immunohistochemical detection of factor VIII/von Willebrand factor antigen (FVIII/vWF-AG) in formalin-fixed paraffin-embedded tissues was investigated using the peroxidase-antiperoxidase (PAP) method. Highly purified human FVIII/vWF was used to raise rabbit anti-FVIII/vWF-AG serum. In addition to anti-FVIII/vWF-AG activity, the unabsorbed antiserum had anti-IgG, anti-IgM, and anti-alpha2-macroglobulin specificities. Following exhaustive absorption with these proteins, the antiserum reacted monospecifically for FVIII/vWF-AG in immunodiffusion, immunoelectrophoresis, and PAP immunohistochemistry. Sections of normal tissues from six patients and a total of 43 neoplasms were examined. Treatment of the tissue sections with trypsin prior to application of the antiserum markedly increased the sensitivity of FVIII/vWF-AG detection. The positive staining for FVIII/vWF-AG was restricted to endothelial cells in both neoplastic and nonneoplastic tissue. In general, the hyperplastic endothelia in neoplastic and reactive tissues stained more intensely than those in normal tissues. Expression of FVIII/vWF-AG by nonendothelial neoplastic cells was not observed. FVIII/vWF-AG is a reliable marker for endothelial cells.

scholarly journals The Expression of hsp-miRNA-200b-3p and -200c-3p in Human Cytomegalovirus-infected Formalin-Fixed, Paraffin-Embedded Tissues

2017 ◽  
Vol 4 (suppl_1) ◽  
pp. S358-S358
Author(s):  
Kyoung Hwa Lee ◽  
Seoyeon Min ◽  
Seul Gi Yoo ◽  
Beom Jin Lim ◽  
Jeong-Hyeon Jo ◽  
...  
Keyword(s):  
Real Time ◽  
Human Cytomegalovirus ◽  
Gene Activation ◽  
Invasive Disease ◽  
Lytic Replication ◽  
Normal Tissues ◽  
Early Protein ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Abstract Background Human cytomegalovirus (HCMV), which exist as asymptomatic latent status, can cause the tissue invasive disease through reactivation in various immunocompromised conditions. Hsp-microRNA has a specific function of post transcriptional suppression through binding with 3’ untranslated region (UTR) of mRNA. In previous study, hsp-miR-200b-3p and -200c-3p had high probability of conjugation with 3’UTR of mRNA encoded by HCMV UL 122–123 region, which translate the immediate early protein 2 (IE2) protein. IE2 (pp86) plays an essential role to initiate and regulate viral early (E) gene activation as well as propagate the subsequent steps of HCMV lytic replication. This study was aimed to evaluate whether HCMV-infected tissue had a lower expression level of hsp-miR-200b-3p and -200c-3p. Methods We had collected the formalin-fixed, paraffin-embedded tissues (FFPEs) with cytopathic pathologic findings as well as positive immunohistochemical stain (IHC) test for HCMV (N = 111). The HCMV-uninfected normal tissues (N = 77) were selected among FFPEs with neither infection nor inflammation as well as negative HCMV IHC test. We performed TaqMan® MicroRNA real-time RT-PCR to measure the expression levels of hsp-miR-200b-3p and -200c-3p and TaqMan® real-time PCR for HCMV UL83 region to measure HCMV viral load in each FFPE. We utilized the standard curves consisting of mirVanaTM miRNA mimics corresponding to each of two miRNAs, ranging from 106 to 101 copies/µL and HCMV NIBSC 09/162 strain, ranging from 5 X 106 to 5 X 101 IU/mL. Results The levels of hsp-miR-200b-3p and -200c-3p were strongly correlated with r=0.844 (P < 0.001). The expressions levels of hsp-miR-200b-3p in HCMV-infected FFPEs (log10 3.50 ± 0.13 copies/µL) were significantly lower than normal tissues (log10 5.24 ± 0.12 copies/µL of input RNA, P < 0.001). Also, HCMV-infected FFPEs were significantly lower levels of hsp-miR-200c-3p compared than normal tissues (log10 5.28 ± 0.18 vs. 7.81 ± 0.11 copies/µL of input RNA, P = 0.025). The levels of miR-200b-3p and -200c-3p had the significant inverse correlation with HCMV VL (200b-3p, spearman r=-0.392, P < 0.001 and 200c-3p, spearman r=-0.355, P < 0.001). Conclusion The low expression of hsp-miRNA-200b-3p and -200c-3p could play a pathophysiologic role of development of HCMV tissue-invasive disease. Disclosures All authors: No reported disclosures.

scholarly journals Automated Extraction of DNA and RNA from a Single Formalin-Fixed Paraffin-Embedded Tissue Section for Analysis of Both Single-Nucleotide Polymorphisms and mRNA Expression

2010 ◽  
Vol 56 (12) ◽  
pp. 1845-1853 ◽  
Author(s):  
Guido Hennig ◽  
Mathias Gehrmann ◽  
Udo Stropp ◽  
Hiltrud Brauch ◽  
Peter Fritz ◽  
...  
Keyword(s):  
Extraction Method ◽  
Nucleotide Polymorphisms ◽  
Automated Extraction ◽  
Single Nucleotide ◽  
Normal Tissues ◽  
Dna And Rna ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Rna And Dna ◽  
Formalin Fixed

BACKGROUND There is an increasing need for the identification of both DNA and RNA biomarkers from pathodiagnostic formalin-fixed paraffin-embedded (FFPE) tissue samples for the exploration of individualized therapy strategies in cancer. We investigated a fully automated, xylene-free nucleic acid extraction method for the simultaneous analysis of RNA and DNA biomarkers related to breast cancer. METHODS We copurified both RNA and DNA from a single 10-μm section of 210 paired samples of FFPE tumor and adjacent normal tissues (1–25 years of archival time) using a fully automated extraction method. Half of the eluate was DNase I digested for mRNA expression analysis performed by using reverse-transcription quantitative PCR for the genes estrogen receptor 1 (ESR1), progesterone receptor (PGR), v-erb-b2 erythroblastic leukemia viral oncogene homolog 2, neuro/glioblastoma derived oncogene homolog (avian) (ERBB2), epoxide hydrolase 1 (EPHX1), baculoviral IAP repeat-containing 5 (BIRC5), matrix metallopeptidase 7 (MMP7), vascular endothelial growth factor A (VEGFA), and topoisomerase (DNA) II alpha 170kDa (TOP2A). The remaining undigested aliquot was used for the analysis of 7 single-nucleotide polymorphisms (SNPs) by MALDI-TOF mass spectrometry. RESULTS In 208 of 210 samples (99.0%) the protocol yielded robust quantification-cycle values for both RNA and DNA normalization. Expression of the 8 breast cancer genes was detected in 81%–100% of tumor tissues and 21%–100% of normal tissues. The 7 SNPs were successfully genotyped in 91%–97% of tumor and 94%–97% of normal tissues. Allele concordance between tumor and normal tissue was 98.9%–99.5%. CONCLUSIONS This fully automated process allowed an efficient simultaneous extraction of both RNA and DNA from a single FFPE section and subsequent dual analysis of selected genes. High gene expression and genotyping detection rates demonstrate the feasibility of molecular profiling from limited archival patient samples.

scholarly journals Formalin-fixed paraffin-embedded renal biopsy tissues: an underexploited biospecimen resource for gene expression profiling in IgA nephropathy

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Sharon Natasha Cox ◽  
Samantha Chiurlia ◽  
Chiara Divella ◽  
Michele Rossini ◽  
Grazia Serino ◽  
...  
Keyword(s):  
Gene Expression ◽  
Iga Nephropathy ◽  
Clinical Decision Making ◽  
Expression Profiles ◽  
Specific Gene ◽  
Renal Lesions ◽  
Formalin Fixed Paraffin ◽  
Renal Biopsies ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Abstract Primary IgA nephropathy (IgAN) diagnosis is based on IgA-dominant glomerular deposits and histological scoring is done on formalin-fixed paraffin embedded tissue (FFPE) sections using the Oxford classification. Our aim was to use this underexploited resource to extract RNA and identify genes that characterize active (endocapillary–extracapillary proliferations) and chronic (tubulo-interstitial) renal lesions in total renal cortex. RNA was extracted from archival FFPE renal biopsies of 52 IgAN patients, 22 non-IgAN and normal renal tissue of 7 kidney living donors (KLD) as controls. Genome-wide gene expression profiles were obtained and biomarker identification was carried out comparing gene expression signatures a subset of IgAN patients with active (N = 8), and chronic (N = 12) renal lesions versus non-IgAN and KLD. Bioinformatic analysis identified transcripts for active (DEFA4,TNFAIP6,FAR2) and chronic (LTB,CXCL6, ITGAX) renal lesions that were validated by RT-PCR and IHC. Finally, two of them (TNFAIP6 for active and CXCL6 for chronic) were confirmed in the urine of an independent cohort of IgAN patients compared with non-IgAN patients and controls. We have integrated transcriptomics with histomorphological scores, identified specific gene expression changes using the invaluable repository of archival renal biopsies and discovered two urinary biomarkers that may be used for specific clinical decision making.

scholarly journals Immunoreactivity of A103, An Antibody to Melan A, in Canine Steroid-Producing Tissues and their Tumors

2001 ◽  
Vol 13 (4) ◽  
pp. 328-332 ◽  
Author(s):  
José A. Ramos-Vara ◽  
Marilyn E. Beissenherz ◽  
Margaret A. Miller ◽  
Gayle C. Johnson ◽  
John M. Kreeger ◽  
...  
Keyword(s):  
Leydig Cells ◽  
Nuclear Staining ◽  
Normal Tissues ◽  
Adrenocortical Adenomas ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Adrenocortical Carcinomas ◽  
Formalin Fixed ◽  
Melanocytic Differentiation ◽  
Sertoli Cell Tumors

The monoclonal antibody A103 to the melanocytic differentiation antigen Melan A stains human steroid-producing cells and their tumors. A total of 200 formalin-fixed, paraffin-embedded canine normal tissues and hyperplastic and neoplastic lesions of the adrenal gland, testis, and ovary were immunohistochemically tested for Melan A with antibody A103. Leydig cell tumors (23/23, 100%), Sertoli cell tumors (14/15, 93%), and adrenocortical adenomas (12/13, 92%) were consistently positive. Adrenocortical carcinomas (23/35, 65%) and granulosa cell tumors (10/17, 59%) were less frequently positive. All pheochromocytomas, seminomas, and dysgerminomas were negative. The pattern of staining was cytoplasmic, but nuclear staining was also frequently seen in normal Leydig cells and their tumors. As in human tumors, immunohistochemistry for Melan A stains many canine steroid-producing tumors and can be used to distinguish these tumors from those of nonstereidogenic cells.

scholarly journals Oncogenic Mutations in Histologically Normal Endometrium: The New Normal?

2019 ◽  
Author(s):  
Vivian Lac ◽  
Tayyebeh M. Nazeran ◽  
Basile Tessier-Cloutier ◽  
Rosalia Aguirre-Hernandez ◽  
Arianne Albert ◽  
...  
Keyword(s):  
Normal Skin ◽  
Digital Pcr ◽  
Driver Mutations ◽  
Normal Endometrium ◽  
Pten Loss ◽  
Normal Tissues ◽  
Oncogenic Mutations ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Tissue Specimens

AbstractThe presence of somatic driver mutations in endometriosis has previously been believed to represent early events in transformation, however our group and others have described such mutations in roughly one-third of cases of deep infiltrating or iatrogenic endometriosis. These forms of endometriosis rarely progress to malignancy. Recent studies have also shown somatic driver mutations in normal skin, blood, peritoneal washings, and esophageal epithelium. Such findings prompt speculation on whether such mutations exist in the eutopic endometrium – the likely tissue of origin of endometriosis. In the current study we investigated the presence of somatic driver mutations in histologically normal endometrium from women lacking evidence of gynecologic malignancy or endometrial hyperplasia. Twenty-five women who underwent hysterectomies and 85 women who underwent endometrial biopsies were included in this study. Formalin-fixed, paraffin-embedded tissue specimens were analyzed by means of targeted sequencing followed by orthogonal validation with droplet digital PCR. PTEN and ARID1A immunohistochemistry (IHC) was also performed as surrogates for inactivating mutations in the respective genes. Overall, we observed somatic driver-like events in over 50% of histologically normal endometrial samples analyzed, which included hotspot mutations in KRAS, PIK3CA, and FGFR2 as well as PTEN-loss by IHC. Analysis of anterior and posterior samplings collected from women who underwent hysterectomies is consistent with the presence of somatic driver mutations within clonal pockets spread throughout the uterus. The prevalence of such oncogenic mutations also increased with age (OR: 1.05 (95% CI: 1.00 – 1.10), p = 0.035). These findings have implications on our understanding of aging and so-called “normal tissues”, thereby necessitating caution in the utilization of mutation-based early detection tools for endometrial or other cancers.

scholarly journals Immunohistochemical Detection of Melanoma-associated Antigens on Formalin-fixed, Paraffin-embedded Canine Tumors

1994 ◽  
Vol 31 (4) ◽  
pp. 455-461 ◽  
Author(s):  
A. J. Berrington ◽  
K. Jimbow ◽  
D. M. Haines
Keyword(s):  
Monoclonal Antibodies ◽  
Periodic Acid ◽  
Specific Antigen ◽  
Periodic Acid Schiff ◽  
Cytoplasmic Staining ◽  
Normal Tissues ◽  
Melanocytic Tumors ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Seven monoclonal antibodies (Moabs), recognizing melanoma-associated antigens in human tissues, were evaluated for their ability to immunohistochemically stain formalin-fixed, paraffin-embedded canine melanomas. Only 2 Moabs, designated human melanosome-specific antigen (HMSA)-1 and HMSA-5, stained routinely processed canine melanomas, staining 21/35 (60%) and 24/35 (69%), respectively. Twenty-nine of 35 (83%) melanomas tested were stained if results of the 2 Moabs were combined. Monoclonal antibody HMSA-1 also stained neoplastic cells of 10/35 (29%) tumors of nonmelanocytic origin and some neurons and salivary gland epithelial cells in normal canine tissues. However, Moab HMSA-1 staining in the nonmelanocytic tumors, consisting of small, discrete periodic acid-Schiff-positivc cytoplasmic droplets, was readily distinguishable from the diffusely granular, cytoplasmic staining of melanocytic tumors. In addition to melanomas, Moab HMSA-5 stained melanocytes and some melanin-containing tumor cells of a pigmented basal cell tumor and melanocytes in normal canine skin. Monoclonal antibodies HMSA-1 and HMSA-5 immunohistochemically identified the majority of canine melanomas, had limited and distinguishable staining in normal tissues and nonmelanocytic tumors, and therefore may be a useful adjunct for the diagnosis of canine melanoma in formalin-fixed, paraffin-embedded tissues.

Antigen Expression in Normal and Neoplastic Canine Tissues Defined by a Monoclonal Antibody Generated against Canine Mesothelioma Cells

1994 ◽  
Vol 31 (6) ◽  
pp. 663-673 ◽  
Author(s):  
K. X. Liu ◽  
A. E. Church Bird ◽  
S. D. Lenz ◽  
S. P. McDonough ◽  
L. G. Wolfe
Keyword(s):  
Monoclonal Antibody ◽  
Smooth Muscle ◽  
Soft Tissues ◽  
Mesothelial Cell ◽  
Bladder Smooth Muscle ◽  
Normal Tissues ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Abc Method ◽  
Formalin Fixed

Monoclonal antibody (MAb) 3B5 generated against canine mesothelioma cells was applied to canine tumors and normal tissues via immunohistochemical and immunoblotting techniques to evaluate antigen binding. By use of an avidin-biotin immunoperoxidase complex (ABC) method, immunoreactivity was noted in reactive mesothelial cells and in normal tissues was observed primarily in mesothelial cell linings, endothelial cells, and smooth muscle of blood vessels and soft tissues; the reactivity was nearly equivalent in frozen or formalin-fixed, paraffin-embedded tissue sections. Use of the ABC method on formalin-fixed, paraffin-embedded tumors yielded moderate to strong cytoplasmic immunostaining of neoplastic cells in 10/11 (91%) mesotheliomas, 18/23 (78%) hemangiosarcomas, 4/10 (40%) intestinal and lung carcinomas, and ≤20% of hemangiomas, leiomyosarcomas, leiomyomas, mammary carcinomas, and squamous cell carcinomas. No immunostaining of tumor cells was observed in fibrosarcomas, hemangiopericytomas, perianal gland carcinomas, and melanomas. Immunoblotting was performed on samples that demonstrated strong immunoreactivity with MAb 3B5 by the ABC method: mesothelioma, hemangiosarcoma, urinary bladder (smooth muscle), and lung (alveolar capillaries). These analyses showed that MAb 3B5 bound a major antigen of 78 kilodaltons (kd) and minor antigens at 56 and 54 kd in normal and neoplastic tissues. The preliminary immunohistochemical results suggest that MAb 3B5 may possess utility in diagnosis of mesotheliomas and hemangiosarcomas, discrimination of cell types in proliferative serosal lesions, and demonstration of vascularity or angiogenesis in neoplastic and inflammatory lesions.

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