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Discrimination of grade 2 and 3 cervical intraepithelial neoplasia by means of analysis of water soluble proteins recovered from cervical biopsies.

Uleberg KE, Munk AC, Brede C, Gudlaugsson E, van Diermen B, Skaland I, Malpica A, Janssen EA, Hjelle A, Baak JP - Proteome Sci (2011)

Bottom Line: The identified proteins are annotated to metabolic processes (36%), signal transduction (27%), cell cycle processes (15%) and trafficking/transport (9%).Using binary logistic regression, Cytokeratin 2 was found to have the strongest independent discriminatory power resulting in 90% overall correct classification. 114 proteins were identified in supernatants from fresh cervical biopsies and many differed between CIN2 and 3.Cytokeratin 2 is the strongest discriminator with 90% overall correct classifications.

View Article: PubMed Central - HTML - PubMed

Affiliation: Pathology Department, Stavanger University Hospital, Armauer Hansen Road 20, Stavanger, Norway. jpabaak@yahoo.com.

ABSTRACT

Background: Cervical intraepithelial neoplasia (CIN) grades 2 and 3 are usually grouped and treated in the same way as "high grade", in spite of their different risk to cancer progression and spontaneous regression rates. CIN2-3 is usually diagnosed in formaldehyde-fixed paraffin embedded (FFPE) punch biopsies. This procedure virtually eliminates the availability of water-soluble proteins which could have diagnostic and prognostic value.

Aim: To investigate whether a water-soluble protein-saving biopsy processing method followed by a proteomic analysis of supernatant samples using LC-MS/MS (LTQ Orbitrap) can be used to distinguish between CIN2 and CIN3.

Methods: Fresh cervical punch biopsies from 20 women were incubated in RPMI1640 medium for 24 hours at 4°C for protein extraction and subsequently subjected to standard FFPE processing. P16 and Ki67-supported histologic consensus review CIN grade (CIN2, n = 10, CIN3, n = 10) was assessed by independent gynecological pathologists. The biopsy supernatants were depleted of 7 high abundance proteins prior to uni-dimensional LC-MS/MS analysis for protein identifications.

Results: The age of the patients ranged from 25-40 years (median 29.7), and mean protein concentration was 0.81 mg/ml (range 0.55 - 1.14). After application of multistep identification criteria, 114 proteins were identified, including proteins like vimentin, actin, transthyretin, apolipoprotein A-1, Heat Shock protein beta 1, vitamin D binding protein and different cytokeratins. The identified proteins are annotated to metabolic processes (36%), signal transduction (27%), cell cycle processes (15%) and trafficking/transport (9%). Using binary logistic regression, Cytokeratin 2 was found to have the strongest independent discriminatory power resulting in 90% overall correct classification.

Conclusions: 114 proteins were identified in supernatants from fresh cervical biopsies and many differed between CIN2 and 3. Cytokeratin 2 is the strongest discriminator with 90% overall correct classifications.

No MeSH data available.


Related in: MedlinePlus

MS2 spectrum of the peptide with the sequence VDLLNQEIEFLK (A), the sequence with the y- and b-series shown (B) and the part of the sequence containing this peptide shown in a multialignment of all identified cytokeratins.
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Figure 5: MS2 spectrum of the peptide with the sequence VDLLNQEIEFLK (A), the sequence with the y- and b-series shown (B) and the part of the sequence containing this peptide shown in a multialignment of all identified cytokeratins.

Mentions: Cytokeratin 2 was identified with five peptides. An MS2 spectrum for one of these peptides is shown in figure 5 panel A. Panel B in the figure show the y- and b-series with the peptide sequence, and panel C a multi-alignment of this part of the protein sequence with all the other cytokeratins identified in this study. The multi-alignment shows that this peptide makes Cytokeratin 2 (CK-2) different from the other cytokeratins identified.


Discrimination of grade 2 and 3 cervical intraepithelial neoplasia by means of analysis of water soluble proteins recovered from cervical biopsies.

Uleberg KE, Munk AC, Brede C, Gudlaugsson E, van Diermen B, Skaland I, Malpica A, Janssen EA, Hjelle A, Baak JP - Proteome Sci (2011)

MS2 spectrum of the peptide with the sequence VDLLNQEIEFLK (A), the sequence with the y- and b-series shown (B) and the part of the sequence containing this peptide shown in a multialignment of all identified cytokeratins.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3142202&req=5

Figure 5: MS2 spectrum of the peptide with the sequence VDLLNQEIEFLK (A), the sequence with the y- and b-series shown (B) and the part of the sequence containing this peptide shown in a multialignment of all identified cytokeratins.
Mentions: Cytokeratin 2 was identified with five peptides. An MS2 spectrum for one of these peptides is shown in figure 5 panel A. Panel B in the figure show the y- and b-series with the peptide sequence, and panel C a multi-alignment of this part of the protein sequence with all the other cytokeratins identified in this study. The multi-alignment shows that this peptide makes Cytokeratin 2 (CK-2) different from the other cytokeratins identified.

Bottom Line: The identified proteins are annotated to metabolic processes (36%), signal transduction (27%), cell cycle processes (15%) and trafficking/transport (9%).Using binary logistic regression, Cytokeratin 2 was found to have the strongest independent discriminatory power resulting in 90% overall correct classification. 114 proteins were identified in supernatants from fresh cervical biopsies and many differed between CIN2 and 3.Cytokeratin 2 is the strongest discriminator with 90% overall correct classifications.

View Article: PubMed Central - HTML - PubMed

Affiliation: Pathology Department, Stavanger University Hospital, Armauer Hansen Road 20, Stavanger, Norway. jpabaak@yahoo.com.

ABSTRACT

Background: Cervical intraepithelial neoplasia (CIN) grades 2 and 3 are usually grouped and treated in the same way as "high grade", in spite of their different risk to cancer progression and spontaneous regression rates. CIN2-3 is usually diagnosed in formaldehyde-fixed paraffin embedded (FFPE) punch biopsies. This procedure virtually eliminates the availability of water-soluble proteins which could have diagnostic and prognostic value.

Aim: To investigate whether a water-soluble protein-saving biopsy processing method followed by a proteomic analysis of supernatant samples using LC-MS/MS (LTQ Orbitrap) can be used to distinguish between CIN2 and CIN3.

Methods: Fresh cervical punch biopsies from 20 women were incubated in RPMI1640 medium for 24 hours at 4°C for protein extraction and subsequently subjected to standard FFPE processing. P16 and Ki67-supported histologic consensus review CIN grade (CIN2, n = 10, CIN3, n = 10) was assessed by independent gynecological pathologists. The biopsy supernatants were depleted of 7 high abundance proteins prior to uni-dimensional LC-MS/MS analysis for protein identifications.

Results: The age of the patients ranged from 25-40 years (median 29.7), and mean protein concentration was 0.81 mg/ml (range 0.55 - 1.14). After application of multistep identification criteria, 114 proteins were identified, including proteins like vimentin, actin, transthyretin, apolipoprotein A-1, Heat Shock protein beta 1, vitamin D binding protein and different cytokeratins. The identified proteins are annotated to metabolic processes (36%), signal transduction (27%), cell cycle processes (15%) and trafficking/transport (9%). Using binary logistic regression, Cytokeratin 2 was found to have the strongest independent discriminatory power resulting in 90% overall correct classification.

Conclusions: 114 proteins were identified in supernatants from fresh cervical biopsies and many differed between CIN2 and 3. Cytokeratin 2 is the strongest discriminator with 90% overall correct classifications.

No MeSH data available.


Related in: MedlinePlus