flavin-adenine-dinucleotide and Uterine-Cervical-Dysplasia

While metabolic changes are considered a cancer hallmark, their assessment has not been incorporated in the detection of early or precancers, when treatment is most effective. Here, we demonstrate that metabolic changes are detected in freshly excised human cervical precancerous tissues using label-free, non-destructive imaging of the entire epithelium. The images rely on two-photon excited fluorescence from two metabolic co-enzymes, NAD(P)H and FAD, and have micron-level resolution, enabling sensitive assessments of the redox ratio and mitochondrial fragmentation, which yield metrics of metabolic function and heterogeneity. Simultaneous characterization of morphological features, such as the depth-dependent variation of the nuclear:cytoplasmic ratio, is demonstrated. Multi-parametric analysis combining several metabolic metrics with morphological ones enhances significantly the diagnostic accuracy of identifying high-grade squamous intraepithelial lesions. Our results motivate the translation of such functional metabolic imaging to

Topics: Cervix Uteri; Epithelium; Female; Flavin-Adenine Dinucleotide; Humans; Metabolic Networks and Pathways; Mitochondrial Dynamics; NAD; NADP; Optical Imaging; Precancerous Conditions; Reproducibility of Results; Sensitivity and Specificity; Staining and Labeling; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms

We report the ex vivo results of an in-house fabricated portable device based on polarized fluorescence measurements in the clinical environment. This device measures the polarized fluorescence and elastic scattering spectra with 405-nm laser and white light sources, respectively. The dominating fluorophore with 405-nm excitation is flavin adenine dinucleotide (FAD) with a fluorescence peak around 510 nm. The measured spectra are highly modulated by the interplay of scattering and absorption effects. Due to this, valuable information gets masked. To reduce these effects, intrinsic fluorescence was extracted by normalizing polarized fluorescence spectra with polarized elastic scattering spectra obtained. A number of fluorophores contribute to the fluorescence spectra and need to be decoupled to understand their roles in the progression of cancer. Nelder-Mead method has been utilized to fit the spectral profile with Gaussian to decouple the different bands of contributing fluorophores (FAD and porphyrin). The change in concentration of FAD during disease progression manifests in the change in ratio of total area to FWHM of its Gaussian profile. Receiver operating characteristic (ROC) curve analysis has been used to discriminate different grades of cervical precancer by using the ratio as input parameter. The sensitivity and specificity for discrimination of normal samples from CIN I (cervical intraepithelial neoplasia) are 75% and 54%, respectively. Further, the normal samples can be discriminated from CIN II samples with 100% and 82% sensitivity and specificity, respectively, and the CIN I from CIN II samples can also be discriminated with 100% sensitivity and 90% specificity, respectively. The results show that the change in the concentration of (FAD) can be used as a marker to discriminate the different grades of the cancer and biochemical changes at an early stage of the cancer can also be monitored with this technique.

Topics: Biomarkers, Tumor; Early Detection of Cancer; Equipment Design; Female; Flavin-Adenine Dinucleotide; Humans; Optical Imaging; Sensitivity and Specificity; Spectrometry, Fluorescence; Uterine Cervical Dysplasia; Uterine Cervical Neoplasms