Anji Anura

Epithelio-mesenchymal transitional attributes in oral sub-mucous fibrosis.

Evaluating molecular attributes in association with its epithelial and sub-epithelial changes of oral sub-mucous fibrosis is meaningful in exploring the plausibility of an epithelio-mesenchymal transition (EMT) and malignant potentiality of this pathosis. In this study histopathological and histochemical attributes for basement membrane and connective tissue in biopsies of oral sub-mucous fibrosis (n = 55) and normal oral mucosa (n = 16) were assessed and expressions of p63, E-cadherin, β-catenin, N-cadherin and TWIST were analyzed immunohistochemically. The p63 and its isoforms (TA and ∆N), PARD3, E-cadherin and β-catenin were also assessed transcriptomically by q-PCR and EMT players like TWIST1, ZEB1, MMP9 and micro-RNA 205 were searched in gene expression microarrays. Oral epithelium demonstrating impairment in progressive maturation in oral sub-mucous fibrosis concomitantly experienced an increase in basement membrane thickness and collagen deposition along with alteration in target molecular expressions. In comparison to non-dysplastic conditions dysplastic stages exhibited significant increase in p63 and p63∆N expressions whereas, E-cadherin and β-catenin exhibited loss from the membrane with concurrent increase in cytoplasm. Further the N-cadherin and TWIST were gained remarkably along with the appearance of nuclear accumulation features of β-catenin. The microarray search had noticed the up-regulation of TWIST1, ZEB1 and MMP9 along with down regulation of micro-RNA 205. The simultaneous increase in basement membrane thickness and sub-epithelial collagen deposition were the plausible indicators for increased matrix stiffness with expected impact on oral epithelial functional homoeostasis. This was corroborated with the increase in expressions of epithelial master regulator p63 and its oncogenic isoform (∆N) along with membranous loss of E-cadherin (EMT hallmark) and its associate β-catein and gain of mesenchymal markers like N-cadherin and TWIST. These also became indicative for the induction of epithelial to mesenchymal transitional mechanism in oral sub-mucous fibrosis when connoted here with the relevant modulation in expressions of EMT regulators.

Computer-aided molecular pathology interpretation in exploring prospective markers for oral submucous fibrosis progression

Evaluation of molecular pathology markers using a computer‐aided quantitative assessment framework would help to assess the altered states of cellular proliferation, hypoxia, and neoangiogenesis in oral submucous fibrosis and could improve diagnostic interpretation in gauging its malignant potentiality.

Computational analysis of p63+ nuclei distribution pattern by graph theoretic approach in an oral pre-cancer (sub-mucous fibrosis)

Background: Oral submucous fibrosis (OSF) is a pre-cancerous condition with features of chronic, inflammatory and progressive sub-epithelial fibrotic disorder of the buccal mucosa. In this study, malignant potentiality of OSF has been assessed by quantification of immunohistochemical expression of epithelial prime regulator-p63 molecule in correlation to its malignant (oral squamous cell carcinoma [OSCC] and normal counterpart [normal oral mucosa [NOM]). Attributes of spatial extent and distribution of p63 + expression in the epithelium have been investigated. Further, a correlated assessment of histopathological attributes inferred from H&E staining and their mathematical counterparts (molecular pathology of p63) have been proposed. The suggested analytical framework envisaged standardization of the immunohistochemistry evaluation procedure for the molecular marker, using computer-aided image analysis, toward enhancing its prognostic value. Subjects and Methods: In histopathologically confirmed OSF, OSCC and NOM tissue sections, p63 + nuclei were localized and segmented by identifying regional maxima in plateau-like intensity spatial profiles of nuclei. The clustered nuclei were localized and segmented by identifying concave points in the morphometry and by marker-controlled watersheds. Voronoi tessellations were constructed around nuclei centroids and mean values of spatial-relation metrics such as tessellation area, tessellation perimeter, roundness factor and disorder of the area were extracted. Morphology and extent of expression are characterized by area, diameter, perimeter, compactness, eccentricity and density, fraction of p63 + expression and expression distance of p63 + nuclei. Results: Correlative framework between histopathological features characterizing malignant potentiality and their quantitative p63 counterparts was developed. Statistical analyses of mathematical trends were evaluated between different biologically relevant combinations: (i) NOM to oral submucous fibrosis without dysplasia (OSFWT) (ii) NOM to oral submucous fibrosis with dysplasia (OSFWD) (iii) OSFWT-OSFWD (iv) OSFWD-OSCC. Significant histopathogical correlates and their corroborative mathematical features, inferred from p63 staining, were also investigated into. Conclusion: Quantitative assessment and correlative analysis identified mathematical features related to hyperplasia, cellular stratification, differentiation and maturation, shape and size, nuclear crowding and nucleocytoplasmic ratio. It is envisaged that this approach for analyzing the p63 expression and its distribution pattern may help to establish it as a quantitative bio-marker to predict the malignant potentiality and progression. The proposed work would be a value addition to the gold standard by incorporating an observer-independent framework for the associated molecular pathology.

Correlated analysis of semi-quantitative immunohistochemical features of E-cadherin, VEGF and CD105 in assessing malignant potentiality of oral submucous fibrosis.

Oral submucous fibrosis, a potentially premalignant condition for oral squamous cell carcinoma, manifests both non-dysplastic and dysplastic grades. Early and specific identification of its malignant potentiality suffers from diagnostic limitations that may be addressed by correlated molecular pathology attributes having histopathological backdrop. Present study correlates expressional alteration in prime epithelial marker E-cadherin, with neo-angiogenic molecules viz. VEGF and CD105 for elucidation of malignant potentiality in different stages of oral submucous fibrosis. Sixty-eight incision biopsies from normal oral mucosa (n = 10), non-dysplastic (n = 18) and different dysplastic grades (n = 40) of oral submucous fibrosis were semi-quantitatively analyzed for immunohistochemical expressions of E-cadherin (membranous and cytoplasmic), VEGF and CD105 which were further statistically correlated. The loss of membranous E-cadherin with increase in cytoplasmic accumulation in differentiative layers of epithelium through the progression of dysplasia was noted along with up-regulation in VEGF expressions. The number of CD105(+) blood vessels and their major axis also showed significant increase from non-dysplasia toward higher grades of dysplasia. The positive correlation between deregulated expression of epithelial cell-cell adhesion molecule and increase in neo-angiogenic attributes of oral submucous fibrosis with increase in dysplastic grades indicated elucidatory potential of molecular expression features in assessment of malignant potentiality in oral submucous fibrosis.

Global spectral and local molecular connects for optical coherence tomography features to classify oral lesions towards unravelling quantitative imaging biomarkers

Correction for ‘Global spectral and local molecular connects for optical coherence tomography features to classify oral lesions towards unravelling quantitative imaging biomarkers’ by Satarupa Banerjee et al., RSC Adv., 2016, 6, 7511–7520.

Identifying dense subgraphs in protein–protein interaction network for gene selection from microarray data

Selection of important genes responsible for a disease is an important task in bioinformatics. Microarray data are often used with differential expression being considered as a cue. Recently, such expression data are supplemented by gene ontology and genes/proteins interaction network for the selection task. The functional knowledge and interaction structure have become critical for understanding the biological processes, including selection of genes potentially associated to complex diseases. In this paper, we propose an approach that combines expression analysis with structural analysis of protein–protein interaction networks to identify genes associated with complex diseases. The dense subgraph structures embedded in the networks are extracted. We present results on three different types of benchmark cancer dataset (prostate cancer, interstitial lung disease and chronic lymphocytic leukemia) and show that several interesting biological information could be inferred, besides achieving a high prediction accuracy. The proposed methodology helps to identify not just differentially expressed genes but also hub genes important in biological processes.

Transfer learning of tissue photon interaction in optical coherence tomography towardsin vivo histology of the oral mucosa.

Oral cancer evolves from different premalignant conditions and the key to save lives is through diagnosis of early symptoms. The conventional practice of post biopsy histopathology reporting is dependent on specificity of sampling site and optical coherence tomography (OCT) imaging is clinically used for guidance. Clinicians infer the tissue constitution by interpreting intensity images and are challenged by inter-and intra-observer variability. In this paper we propose transfer learning of tissue specific photon interaction statistical physics in swept-source OCT for characterizing the oral mucosa with the aim of reducing this reporting variability. The source task models statistical physics of ballistic and near-ballistic photons and its intensity attenuation and target task learns the parameters obtained by solving the source task to identify co-located heterogeneity of tissues. Performance is compared with conventional histopathology of healthy, premalignant and malignant oral lesions supporting its use towards in vivo histology of the oral mucosa for pre-biopsy screening.

Nanomechanical signatures of oral submucous fibrosis in sub-epithelial connective tissue

Oral sub-mucous fibrosis (OSF), a potentially malignant disorder, exhibits extensive remodeling of extra-cellular matrix in the form of sub-epithelial fibrosis which is a possible sequel of assaults from different oral habit related irritants. It has been assumed that micro/nanobio-mechanical imbalance experienced in the oral mucosa due to fibrosis may be deterministic for malignant potential (7-13%) of this pathosis. Present study explores changes in mechanobiological attributes of sub-epithelial connective tissue of OSF and the normal counterpart. The atomic force microscopy was employed to investigate tissue topography at micro/nano levels. It documented the presence of closely packed parallel arrangement of dense collagen fibers with wide variation in bandwidth and loss of D-space in OSF as compared to normal. The AFM based indentation revealed that sub-epithelium of OSF tissue has lost its flexibility with increased Youngs modulus, stiffness, adhesiveness and reduced deformation of the juxta-epithealial connective tissue towards the deeper layer. These significant variations in nano-mechanical properties of the connective tissue indicated plausible impacts on patho-physiological microenvironment. Excessive deposition of collagen I and diminished expression of collagen III, fibronectin along with presence of α-SMA positive myofibroblast in OSF depicted its pathological basis and indicated the influence of altered ECM on this pathosis. The mechanobiological changes in OSF were corroborative with change in collagen composition recorded through immunohistochemistry and RT-PCR. The revelation of comparative nanomechanical profiles of normal oral mucosa and OSF in the backdrop of their structural and cardinal molecular attributes thus became pivotal for developing holistic pathobiological insight about possible connects for malignant transformation of this pre-cancer.

Endorsing cellular competitiveness in aberrant epithelium of oral submucous fibrosis progression: neighbourhood analysis of immunohistochemical attributes

Epithelial abnormality during the transformation of oral submucous fibrosis (OSF) into oral squamous cell carcinoma has been well studied and documented. However, the differential contribution of atrophy and hyperplasia for malignant potentiality of OSF is yet to be resolved. Existing diagnostic conjectures lack precise diagnostic attributes which may be effectively resolved by substantiation of specific molecular pathology signatures. Present study elucidates existence of cellular competitiveness in OSF conditions using computer-assisted neighbourhood analysis in quantitative immunohistochemistry (IHC) framework. The concept of field cancerization was contributory in finding correspondence among neighbouring cells of epithelial layers with reference to differential expression of cardinal cancer-related genes [c-Myc (oncogene), p53 (tumour suppressor), and HIF-1α (hypoxia regulator)] which are known to be important sensors in recognizing cellular competitive interface. Our analyses indicate that different states of OSF condition may be associated with different forms of competitiveness within epithelial neighbouring cells which might be responsible to shape the present and future of the pre-malignant condition. Analytical findings indicated association of atrophic epithelium with stress-driven competitive environment having low c-Myc, high-p53, and stable HIF-1α (the looser cells) which undergo apoptosis. Whereas, the cells with high c-Myc+ (winner cells) give rise to hyperplastic epithelium via possible mutation in p53. The epithelial dysplasia plausibly occurs due to clonal expansion of c-Myc and p53 positive supercompetitor cells. Present study proposes quantitative IHC along with neighbourhood analysis which might help us to dig deeper on to the interaction among epithelial cell population to provide a better understanding of field cancerization and malignant transformation of pre-malignancy.