Nuclear Morphometry Research Articles

Assessment of (EZH2) expression, digital image analysis of CD8+ lymphocytes, and Automated nuclear morphometry in renal cell carcinoma

Assessment of (EZH2) expression, digital image analysis of CD8+ lymphocytes, and Automated nuclear morphometry in renal cell carcinoma

Machine Learning Streamlines the Morphometric Characterization and Multiclass Segmentation of Nuclei in Different Follicular Thyroid Lesions: Everything in a NUTSHELL

The diagnostic assessment of thyroid nodules is hampered by the persistence of uncertainty in borderline cases and further complicated by the inclusion of noninvasive follicular tumor with papillary-like nuclear features (NIFTP) as a less aggressive alternative to papillary thyroid carcinoma (PTC). In this setting, computational methods might facilitate the diagnostic process by unmasking key nuclear characteristics of NIFTP. The main aims of this work were to (1) identify morphometric features of NIFTP and PTC that are interpretable for the human eye and (2) develop a deep learning model for multiclass segmentation as a support tool to reduce diagnostic variability. Our findings confirmed that nuclei in NIFTP and PTC share multiple characteristics, setting them apart from hyperplastic nodules (HP). The morphometric analysis identified 15 features that can be translated into nuclear alterations readily understandable by pathologists, such as a remarkable internuclear homogeneity for HP in contrast to a major complexity in the chromatin texture of NIFTP and to the peculiar pattern of nuclear texture variability of PTC. A few NIFTP cases with available next-generation sequencing data were also analyzed to initially explore the impact of RAS-related mutations on nuclear morphometry. Finally, a pixel-based deep learning model was trained and tested on whole-slide images of NIFTP, PTC, and HP cases. The model, named NUTSHELL (NUclei from Thyroid tumors Segmentation to Highlight Encapsulated Low-malignant Lesions), successfully detected and classified the majority of nuclei in all whole-slide image tiles, showing comparable results with already well-established pathology nuclear scores. NUTSHELL provides an immediate overview of NIFTP areas and can be used to detect microfoci of PTC within extensive glandular samples or identify lymph node metastases. NUTSHELL can be run inside WSInfer with an easy rendering in QuPath, thus facilitating the democratization of digital pathology.

O-307 Decrease of human sperm hypercondensed chromatin volume after slow freezing revealed by three-dimensional (3D) nuclear morphometry

Abstract Study question Are the 3D morphometry parameters of the human sperm nucleus affected by slow freezing? Summary answer The slow freezing-thawing cycle induces significant alterations on hypercondensed nucleus zone by reducing its volume and splits it into several small zones. What is known already Sperm cryopreservation is an essential technique for infertility care, but it seems to highly affect the genetic and epigenetic integrity of sperm nuclei with poorly identified mechanisms. Moreover, it is known that sperm nuclear alterations may be involved in pregnancy failures. Therefore, sperm nuclear biomarkers are an essential issue to assess the outcome of ART procedures when frozen sperm is used. Thereby, 3D visualization of chromatin is a good way to understand modifications in the organization and composition of sperm chromatin after a slow freezing procedure. Study design, size, duration Ten semen of patients were analyzed (Germetheque biobank) after informed consent. Standard semen parameters, DNA fragmentation, chromatin decondensation, sperm telomere length (STL) and 3D nuclear morphometry were performed before and after slow freezing. Morphometric measurements were performed from a digital representation of the sperm nucleus in 3 dimensions that allow us to measure the volume, flatness and elongation of each sperm nucleus and characterize hypercondensed zones, with live/dead spermatozoa distinction. Participants/materials, setting, methods After addition of cryoprotective medium v/v (Cryosperm®), sperm was frozen with a programmable freezer (NanoDigitcool®). Standards sperm parameters were measured in accordance with WHO guidelines 2021, DNA fragmentation by flow cytometry TUNEL assay, chromatin decondensation by epifluorescent microscopy with Chromomycin A3 and STL using qPCR. 3D nuclear representation was performed by a z-axis image acquisition series of DAPI/PI stained slides with optigrid epifluorescent microscopy. Analyses were carried out using the NucleusJ2.0 software with NODeJ plugin. Main results and the role of chance Freezing induces an alteration of sperm vitality (p < 0.0001) and progressive and total motility (p < 0.001 and p < 0.0001 respectively) and an increase in the percentage of sperm with DNA fragmentation (18.30% fresh vs 31.6% frozen, p < 0.05). We did not observe a modification of the median volume of the sperm nucleus before and after freezing (37.15 [IC = 36.5;38.14] and 37.10 [33.27;37.78] µm3 respectively), as the same for flatness and elongation (2.48 vs. 2.50 a.u. and 2.04 vs. 2.04 a.u. respectively). Nevertheless, five samples showed significant alterations of nucleus volume (p < 0.0001), due to diminution of flatness and more or less elongated. A significant decrease of the total volume of the hypercondensed chromatin zone was observed after freezing (0.94 [0.84;0.99] before vs. 0.61 [0.53;0.69] µm3 after freezing-thawing, p < 0.0001), with an effect size of -1.48 [-2.10; -0.86]. Decrease of total volume of hypercondensed region was measured homogeny in all the samples tested, particularly on live (1.00 vs. 0.68 µm3 respectively, p < 0.0001) compared with dead spermatozoa (0.72 vs. 0.57 µm3, p < 0.05). We showed also a significant increase of the number of hypercondensed zones in each nucleus (p < 0.001), reflecting a split into several parts of the hypercondensed chromatin. Limitations, reasons for caution A 3D-FISH analysis would allow to characterize the hypercondensed chromatin portion, probably as heterochromatin and which we suspect to be the nucleus ring. Moreover, a chromosomes specific marking would allow for the identification of whether whole chromosomes or specific parts of chromosomes (centromeres or telomeres) are affected by this decompaction. Wider implications of the findings This study is the first to make exacts measurements of human sperm nucleus because of a non-decondensing method. We demonstrate that slow freezing affects hypercondensed chromatin probably through a decondensation and modification of the nucleus-specific organization. This new assessment will allow a better proficiency of men fertility preservation techniques. Trial registration number NCT04715828

Automated Nuclear Morphometry: A Deep Learning Approach for Prognostication in Canine Pulmonary Carcinoma to Enhance Reproducibility.

The integration of deep learning-based tools into diagnostic workflows is increasingly prevalent due to their efficiency and reproducibility in various settings. We investigated the utility of automated nuclear morphometry for assessing nuclear pleomorphism (NP), a criterion of malignancy in the current grading system in canine pulmonary carcinoma (cPC), and its prognostic implications. We developed a deep learning-based algorithm for evaluating NP (variation in size, i.e., anisokaryosis and/or shape) using a segmentation model. Its performance was evaluated on 46 cPC cases with comprehensive follow-up data regarding its accuracy in nuclear segmentation and its prognostic ability. Its assessment of NP was compared to manual morphometry and established prognostic tests (pathologists' NP estimates (n = 11), mitotic count, histological grading, and TNM-stage). The standard deviation (SD) of the nuclear area, indicative of anisokaryosis, exhibited good discriminatory ability for tumor-specific survival, with an area under the curve (AUC) of 0.80 and a hazard ratio (HR) of 3.38. The algorithm achieved values comparable to manual morphometry. In contrast, the pathologists' estimates of anisokaryosis resulted in HR values ranging from 0.86 to 34.8, with slight inter-observer reproducibility (k = 0.204). Other conventional tests had no significant prognostic value in our study cohort. Fully automated morphometry promises a time-efficient and reproducible assessment of NP with a high prognostic value. Further refinement of the algorithm, particularly to address undersegmentation, and application to a larger study population are required.

Use of topical methylene blue to image nuclear morphometry with a low-cost scanning darkfield microendoscope.

Fiber-optic microendoscopy is a promising approach to noninvasively visualize epithelial nuclear morphometry for early cancer and precancer detection. However, the broader clinical application of this approach is limited by a lack of topical contrast agents available for in vivo use. The aim of this study was to evaluate the ability to image nuclear morphometry in vivo with a novel fiber-optic microendoscope used together with topical application of methylene blue (MB), a dye with FDA approval for use in chromoendoscopy in the gastrointestinal tract. The low-cost, high-resolution microendoscope implements scanning darkfield imaging without complex optomechanical components by leveraging programmable illumination and the rolling shutter of the image sensor. We validate the integration of our system and MB staining for visualizing epithelial cell nuclei by performing ex vivo imaging on fresh animal specimens and in vivo imaging on healthy volunteers. The results indicate that scanning darkfield imaging significantly reduces specular reflection and resolves epithelial nuclei with enhanced image contrast and spatial resolution compared to non-scanning widefield imaging. The image quality of darkfield images with MB staining is comparable to that of fluorescence images with proflavine staining. Our approach enables real-time microscopic evaluation of nuclear patterns and has the potential to be a powerful noninvasive tool for early cancer detection.

Significance of Transmembrane 9 Superfamily 4 (TM9SF4) immunohistochemical expression and nuclear morphometry in premalignant colonic lesions and colorectal carcinoma

Significance of Transmembrane 9 Superfamily 4 (TM9SF4) immunohistochemical expression and nuclear morphometry in premalignant colonic lesions and colorectal carcinoma

Significance of Sex Determining Region Y-Box transcription factor 4 (SOX4) expression and Nuclear morphometry in endometrial carcinoma

Significance of Sex Determining Region Y-Box transcription factor 4 (SOX4) expression and Nuclear morphometry in endometrial carcinoma

Evaluation of nuclear morphometry in exfoliative cytology of buccal mucosa in patients with high risk of oral cancer

BackgroundOral cancer poses a significant global health burden, with India having the highest prevalence. Effective detection is crucial in effective prevention. This study aimed to evaluate nuclear morphometric parameters (NMPs) in buccal mucosa cells of smokers, correlate NMPs with dysplasia, establish cut off values for grading dysplasia, and investigate the relationship between NMPs and smoking. MethodsAfter obtaining ethical approval and informed consent, patients were recruited from the outpatient department of our institution. A target sample size of 250 was calculated. The data included smoking exposure quantified in pack-years, nuclear morphometric analysis (NMA) of buccal mucosa cells obtained through oral cytology using Image J, and the severity of dysplasia of the slides assessed by pathologists. Statistical analysis assessed the impact of dysplasia and the association between nuclear characteristics and smoking exposure. Receiver operating characteristic (ROC) plots determined the potential of these parameters to distinguish dysplasia levels. ResultsSignificant differences in NMPs were observed among different smoking groups. Dysplasia severity had a significant correlation with NMPs, and strong correlations were found between NMPs and lifetime smoking exposure. ROC analysis established cut off values for NMPs with good sensitivity and specificity for classifying dysplasia severity. ConclusionsThis study highlights the potential of NMA as a tool for oral cancer screening. NMPs can distinguish dysplasia severity and correlate with tobacco (smoking). The efficiency of NMA in a non-invasive oral cytology offers promise for patient-centered screening Additionally, the findings suggest future applications in telepathology and the potential for AI integration in automated screening after conducting multicentric large-scale studies.

Nuclear morphometry and expression of CK20, p53 and Ki-67 in the diagnosis of bladder transitional cell carcinomas in dogs

Nuclear morphometry and expression of CK20, p53 and Ki-67 in the diagnosis of bladder transitional cell carcinomas in dogs

Study of nuclear morphometry in the cytologic evaluation of thyroid neoplasms

Study of nuclear morphometry in the cytologic evaluation of thyroid neoplasms

Mule embryos share identical morphological features to horse embryos

This study aimed to compare the morphometry of horse and mule embryos. The study's hypothesis was that the micronuclei and nuclear fragmentation indexes are higher in mule embryos than in horse embryos. Twenty-two mares were randomly assigned in a crossover design to receive semen from a horse and a donkey; thirteen horse and thirteen mule embryos were obtained. Embryos were recovered eight days post-ovulation and classified according to the stage of development and quality with a score from 1 (excellent) to 4 (degenerate). Embryos were stained with Hoechst33342, and images were acquired with a fluorescence microscope. Nuclei were categorized as compact, mitotic, or fragmented; the fragmented and mitotic indexes were calculated based on their proportion over the total amount of nuclei counted. Embryo size and nuclear morphometry were assessed through ImageJ. Data analyses were carried out with GraphPad using ANOVA and T-test; significance was set at P < 0.05. The number of positive flushes in cycles bred with donkey or stallion semen did not differ when compared per cycle or per ovulation (13 vs. 12) (P > 0.05). One set of twins was recovered from a mare bred to the stallion that had a double ovulation; a mule and horse embryos were both recovered from eight mares. There was no difference in size between mule and horse embryos (915.5 ± 288 μm vs. 575.8 ± 69.6 μm) (P > 0.05) size of the study. The mule embryos scored between grade 1 (n = 9) and grade 2 (n = 4); similarly, the horse embryos scored between grade 1 (n = 6) and grade 2 (n = 7). The evaluation of the nuclear morphometry revealed that horse and mule embryos have a similar number of compact nuclei per sector (148.7 ± 6.8 nuclei/sector in mule embryos vs. 156.5 ± 8.5 nuclei/sector in horse embryos) (P > 0.05); however, the number of mitotic nuclei tended to be higher in mule embryos (5.2 ± 0.82) than in horse embryos (3.3 ± 0.3) (P = 0.08). The fragmented nuclei index was similar between mule (0.25 ± 0.1%) and horse (0.22 ± 0.1%) embryos (P = 0.4); the mitotic nuclei index was higher in mule embryos (3.2 ± 0.4%) than in horse embryos (2.2 ± 0.2%) (P = 0.02). In conclusion, embryo morphology of mares bred to a donkey and a horse shares similar nuclear ultrastructure features, except that mule embryos have a higher mitotic index.

Autophagy and nuclear morphometry are associated with histopathologic features in esophageal squamous cell carcinoma.

Less than 15% of patients with esophageal squamous cell carcinoma (ESCC) survive 5years after diagnosis. A better understanding of the biology of these tumors and the development of clinical biomarkers is needed. Autophagy is a physiological mechanism involved in the turnover of cellular components that plays a key role in cancer. This study evaluated the differential levels of three key regulators of autophagy (SQSTM1, MAP1LC3B, and BECN1) in patients with ESCC, associating autophagy with histopathologic features, including the grade of differentiation, mitotic rate, inflammation score, and the intensity of tumor-infiltrating lymphocytes. Nuclear morphometry of the tumor parenchyma was also assessed, associating it with autophagy and histopathology. All three markers significantly increased in patients with ESCC compared to the control group. Based on the mean expression of each protein in the control group, 57% of patients with ESCC had high levels of all three markers compared to control patients (14%). The most frequent profiles found in ESCC were BECNhigh/MAP1LC3high and BECNhigh/SQSTM1high. According to the TCGA database, we found that the main autophagy genes were upregulated in ESCC. Moreover, high levels of autophagy markers were associated with a poor prognosis. Considering nuclear morphometry, ESCC samples showed a significant reduction in nuclear area, which was strongly negatively correlated with autophagy. Finally, the percentage of normal nuclei was associated with tumor differentiation, while poorly differentiated tumors showed lower SQSTM1 levels. ESCC progression may involve increased autophagy and changes in nuclear structure, associated with clinically relevant histopathological features. KEY MESSAGES: Autophagy markers are co-increased in primary ESCC. Autophagy negatively correlates with nuclear morphometry in ESCC parenchyma. Autophagy and nuclear morphometry are associated with histopathological features. Autophagy is increased in ESCC-TCGA database and associated with poor prognosis.

Morphometric study in thyroid tumors.

Various morphonuclear studies using digital image analysis have been taken into account in order to establish the malignancy of thyroid lesions based on their size and on the chromatographic characteristics of tumor cell nuclei. Nuclear morphometry involves the measurement of nuclear parameters to obtain diagnostically important information in an objective and reproducible manner. The aim of the present study was to evaluate the detailed morphometric analysis of histopathological preparations with lesions of the thyroid gland and to investigate its role in differentiating between benign and malignant thyroid lesions. The present study included 10 benign and 26 malignant thyroid cases with different selected thyroid lesions. Using a microscope connected to a computerized video system, nuclear morphometric parameters including the nuclear area, perimeter, average intensity, red average, width and roundness, were measured and analyzed. The main parameters used in the statistical calculation were significant in distinguishing between benign and malignant thyroid lesions. The association of morphometry in cytological smears for suspected malignant follicular lesions led to increased accuracy in establishing a suspicious malignant diagnosis for follicular lesions. Nuclear morphometry provides an unbiased point of view that increases diagnosis accuracy. Computerized morphometry can positively influence diagnostic accuracy, allowing for a better correlation with clinical and imaging data.

Nuclear and chromatin rearrangement associate to epigenome and gene expression changes in a model of in vitro adipogenesis and hypertrophy

Hypertrophy of adipocytes represents the main cause of obesity. We investigated in vitro the changes associated with adipocyte differentiation and hypertrophy focusing on the nuclear morphometry and chromatin epigenetic remodelling. The 3 T3-L1 pre-adipocytes were firstly differentiated into mature adipocytes, then cultured with long-chain fatty acids to induce hypertrophy. Confocal and super-resolution stimulation emission depletion (STED) microscopy combined with ELISA assays allowed us to explore nuclear architecture, chromatin distribution and epigenetic modifications. In each condition, we quantified the triglyceride accumulation, the mRNA expression of adipogenesis and dysfunction markers, the release of five pro-inflammatory cytokines. Confocal microscopy revealed larger volume and less elongated shape of the nuclei in both mature and hypertrophic cells respect to pre-adipocytes, and a trend toward reduced chromatin compaction. Compared to mature adipocytes, the hypertrophic phenotype showed larger triglyceride content, increased PPARγ expression reduced IL-1a release, and up-regulation of a pool of genes markers for adipose tissue dysfunction. Moreover, a remodelling of both epigenome and chromatin organization was observed in hypertrophic adipocytes, with an increase in the average fluorescence of H3K9 acetylated domains in parallel with the increase in KAT2A expression, and a global hypomethylation of DNA. These findings making light on the nuclear changes during adipocyte differentiation and hypertrophy might help the strategies for treating obesity and metabolic complications.

Significance of Nuclear Morphometry in Breast Lesions: A Cross-Sectional Study.

Background Fine-needle aspiration cytology (FNAC) is one of the reliable methods in diagnosing breast cancers. Morphometric studies are done in benign and malignant neoplasms of various organs by using software, which measures cellular, cytoplasmic, and nuclear parameters. Nuclear parameters define the behavior of the neoplasm. This study aims to evaluate nuclear morphometry parameters in aspirated smears of breast lesions and determine the association between cytological findings with nuclear morphometry parameters. Methodology It's a retrospective cytology study from July 2020 to June 2022 conducted at a tertiary health care center in Kolar, Karnataka, India. The FNAC smears of breast mass were analyzed cytologically and were subjected to nuclear morphometry study. Nuclear parameters such asnuclear area, nuclear perimeter, nuclear Feret diameter, minimum Feret, and shape factor were captured inZen software (Zeiss, Oberkochen, Germany) and ImageJ software (National Institutes of Health,Bethesda, MD, USA;Laboratory for Optical and Computational Instrumentation [LOCI], University of Wisconsin-Madison, Madison, WI, USA). The association between nuclear morphometric findings and cytological findings was noted. A descriptive statistical analysis was done. Results Sixty cases of mass in the breast were considered for the study of which37 cases were benign and 23 were malignant. Nuclear morphometry parameters such asnuclear area, nuclear perimeter, nuclear Feret diameter, minimum Feret, and shape factor for benign breast lesions were 25.16 ± 3.2 µm2, 21.58 ± 1.89 µm, 6.5± 0.94 µm, 4.87 ± 0.50 µm, and 0.92± 0.02, respectively, and for malignant breast cases were 46.57 ± 12.24 µm2, 27.53 ± 3.26 µm, 10.08 ± 1.18 µm, 6.49± 0.88 µm, and 0.93 ± 0.01, respectively. The association of all nuclear parameters between benign and malignant lesions was statistically significant (P= 0.001). Conclusions Nuclear morphometric study in breast lesions is a concept that supplements FNAC findings in differentiating benign from malignant lesions.

Nuclear morphometry and stereological analysis of gastrocnemius and pectoralis major muscle tissue from MDX mice supplemented or not with creatine for sixteen weeks

Duchenne muscular dystrophy is a genetic disease that causes a reduction in the expression of dystrophin, thus, with the reduction of the levels of this protein, a progression of muscle atrophy, reduction of strength and physical impairments is identified, which culminates in death from inability to contract the respiratory muscles. Creatine is a dietary supplement that helps maintain strength and recent findings indicate a potential anti-inflammatory effect of this amine. The present study aimed to compare the progress of muscular dystrophy in terms of inflammation, presence of intramuscular glycogen and tissue fibrosis in the gastrocnemius and pectoralis major muscles of MDX mice. We selected 20 MDX mice that were supplemented or not with creatine at a dosage of 0.3 grams per kilogram of body weight, for 16 weeks. After that, the gastrocnemius and pectoralis major were extracted for morphological and morphometric analysis. The inflammatory progress derived from Duchenne muscular dystrophy affects animals, even when supplemented, however, in a less forceful and disabling way, enabling them to have less functional damage. Nevertheless, the supplemented animals have a greater stock of intramuscular glycogen in the evaluated gastrocnemius tissue, a factor that corroborates with the survival and better quality of movement in these animals, given the availability of energy for the contraction of this musculature. Furthermore, a lower frequency and extent of tissue fibrosis is observed in supplemented animals, aspects that enable longer survival and qualify creatine, even in an experimental condition, as an auxiliary supplement in the treatment of a severe dystrophic condition, acting mainly in the maintenance of strength and reduction of local tissue fibrosis.

Quantitative Nuclear Grading: An Objective, Artificial Intelligence–Facilitated Foundation for Grading Noninvasive Papillary Urothelial Carcinoma

In nonmuscle invasive bladder cancer, grade drives important treatment and management decisions. However, grading is complex and qualitative, and it has considerable interobserver and intraobserver variability. Previous literature showed that nuclear features quantitatively differ between bladder cancer grades, but these studies were limited in size and scope. In this study, we aimed to measure morphometric features relevant to grading criteria and build simplified classification models that objectively distinguish between the grades of noninvasive papillary urothelial carcinoma (NPUC). We analyzed 516 low-grade and 125 high-grade 1.0-mm diameter image samples from a cohort of 371 NPUC cases. All images underwent World Health Organization/International Society of Urological Pathology 2004 consensus pathologist grading at our institution that was subsequently validated by expert genitourinary pathologists from 2 additional institutions. Automated software segmented the tissue regions and measured the nuclear features of size, shape, and mitotic rate for millions of nuclei. Then, we analyzed differences between grades and constructed classification models, which had accuracies up to 88% and areas under the curve as high as 0.94. Variation in the nuclear area was the best univariate discriminator and was prioritized, along with the mitotic index, in the top-performing classifiers. Adding shape-related variables improved accuracy further. These findings indicate that nuclear morphometry and automated mitotic figure counts can be used to objectively differentiate between grades of NPUC. Future efforts will adapt the workflow to whole slides and tune grading thresholds to best reflect time to recurrence and progression. Defining these essential quantitative elements of grading has the potential to revolutionize pathologic assessment and provide a starting point from which to improve the prognostic utility of grade.

Optical nanoscopy approach to unveil chromatin alterations implicated in neuroblastoma transformation.

Chromatin organization is a well-orchestrated mechanism regulating cellular physiology and its changes are associated with the onset of many diseases, including cancer. Neuroblastoma (NB) is the most common extracranial tumor in childhood, and it shows remarkable heterogeneity in the phenotypes and evolutionary patterns. Although the molecular mechanisms of this heterogeneity are still unclear, the impairment of the “epigenetic machinery” and the consequent chromatin reorganization might play an important role. We aim to assess the possible alterations in chromatin nanoscale architecture correlated with NB transformation using NB cells genetically engineered to overexpress the non-coding RNA NDM29, which promotes differentiation of tumor cells into a neuronal lineage. In both malignant NB cells (Mock) and neuron-like cells (S1.1), we explored chromatin organization and nuclear architecture by optical nanoscale microscopy using both confocal and stimulation emission depletion (STED) imaging methods, in parallel with molecular biology analyses. We characterized the nuclear 3D morphology of Mock vs. S1.1 cells by measuring volume, shape, and circularity of the nuclei as well as chromatin compaction and distribution. We imaged the immunolabeled histone H3 modifications (H3K9 trimethylation and H3K9 acetylation) to characterize chromatin regions typically associated with highly and poorly condensed chromatin, respectively. In parallel, we quantified the global DNA methylation status by measuring the levels of 5-methylcytosine (5-mC) using the enzyme-linked immunosorbent assay (ELISA) MethylFlash Global DNA. Comparing Mock vs. S1.1 cells, we observed (i) significant remodeling of nuclear morphometry; (ii) marked rearrangement of chromatin domains rich in highly-acetylated and -methylated H3K9; (iii) changes in average chromatin compaction and DNA methylation. Altogether, the results point to an involvement of epigenetic mechanisms which may sustain the chromatin remodeling during NB malignant transformation, paving the way towards new prognostic and therapeutic approaches and understanding chromatin remodeling in physiological/pathological processes.

Evaluation of Nuclear Morphometry and Ki-67 Proliferative Marker in Astrocytomas: A Ambispective Study

Introduction: Astrocytomas are the commonest primary Central Nervous System (CNS) tumours. Its diagnosis is based on histopathological criteria defined by the World Health Organization (WHO) 2016 that grades astrocytoma into four grades. The subjective nature of WHO grading has prompted for more objective methods to evaluate nuclear features. Furthermore, Ki-67, a marker of cellular proliferation is a useful diagnostic tool that also helps in prognostic evaluation and to plan adjuvant therapy in astrocytomas. Aim: To evaluate the nuclear morphometry and Ki-67 proliferative marker in astrocytomas and to assess the relationship of WHO grade with proliferative activity using Ki-67 immunostaining. Materials and Methods: This ambispective study was conducted in the Department of Pathology, JSS Academy of Higher Education and Research, Mysuru, Karnataka, India, with a total duration of four years (October 2017 to September 2021). From 35 astrocytoma cases, Haematoxylin and Eosin (H&amp;E) stained slides were retrieved and reviewed appropriately by two pathologists and graded using the WHO criteria. Nuclear morphometric analysis was performed on the H&amp;E slides using Olympus BX-41 research microscope. The parameters evaluated were mean nuclear length, mean nuclear diameter, mean nuclear perimeter, mean nuclear area, Mean Nuclear Roundness Factor (MNRF) and mean nuclear ellipse form. The cases were then stained with Ki-67 antibody. The relationships between WHO grade of astrocytoma and nuclear morphometry and WHO grade with proliferative index was analysed. Results: According to WHO grading, four cases were Grade 1 (11.4%), 12 grade 2 (34.3%), six grade 3 (17.1%) and 13 grade 4 (37.1%). Significant correlation was seen between WHO grading and mean nuclear length, diameter, perimeter, area and Ki-67 with p-value of less than 0.001. Conclusion: Astrocytoma is an extremely heterogeneous disease with unpredictable outcome. The widely used WHO grading is subjective, while nuclear morphometry, using computer assisted image analysis, can ensure more objective assessment. The Ki-67 index could provide valuable information and may compliment the other parameters.

Significance of Nuclear Morphometry in Salivary Gland Neoplasms: A Cross-sectional Study

Introduction: Fine Needle Aspiration Cytology (FNAC) is one of the commonest and most efficient investigation done for salivary gland swellings, which is of much help in diagnosing inflammatory, benign and malignant lesions. The morphology of nucleus determines the behaviour of the cell/tissue as benign or malignant. Nuclear morphometry analysis is done by using various software. Aim: To determine the nuclear morphometry parameters in salivary gland neoplasm of FNAC smears by Image J software and to determine its association with the cytological diagnosis. Materials and Methods: This was a retrospective laboratory observational cross-sectional study done for four years (2018- 2021) at a tertiary healthcare centre, attached to Sri DevarajUrs Medical College, Kolar, Karnataka, India. All the FNAC smears of salivary gland neoplasms were considered and classified by cytomorphology as benign and malignant tumours. All smears were analysed using Image J software for nuclear morphometric parameters such as area of the nucleus, perimeter of the nucleus, feret diameter, minimum ferret and skewness. The nuclear morphometric findings of benign and malignant tumours were compared and statistical analysis was doneby using Student’s t-test, p-value of &lt;0.05 was considered statistically significant. Results: A total of 52 cases were studied. The average age of presentation of benign and malignant salivary gland neoplasm was 38.7 and 49.4 years, respectively. Among, 30 (57.6%) benign neoplasms there were 23 (76.7%), pleomorphic adenoma was the commonest and among 22 (42.4%) malignant squamous cell carcinoma 16 (72.7%) deposits was the commonest. The nuclear morphometry analysis showed that the mean values of area (p-value ≤0.001), perimeter (p-value≤0.001), ferret diameter (p-value≤0.001) and minimum ferret (p-value=0.001) of malignant lesions were comparatively higher than benign and was statistically significant. Conclusion: Nuclear morphometry along with routine cytopathological evaluation will improve the accuracy of diagnosis of neoplastic lesions of salivary gland. This information can be used to plan better treatment and predict prognosis of the disease. Digital morphological analysis helps in obtaining quantitative values from qualitative data which can be utilised further for automation.