Mitotic Features Research Articles

Abstract 6231: Multi-omic characterisation of clear cell renal carcinoma reveals endogenous and exogenous processes related to its aetiology

Abstract Endogenous and exogenous exposures are associated with distinctive molecular marks in somatic tissues, including human tumours. Integrative multi-omic approaches could help improving the resolution to detect the molecular marks associated with disease aetiology. Here, we leveraged the overlapping molecular information of clear cell renal cell carcinomas (ccRCC) tumours from Mutographs cohort by integrating DNA methylation, transcriptome, and whole-genome sequencing-based somatic mutation data to explore and gain new insights into exposures linked to ccRCC aetiology. Our novel framework identified important sources of biological variance across ccRCC tumours that were summarized as molecular components. We inferred these molecular ccRCC components into independent datasets to further explore their relationship with molecular and epidemiological features of ccRCC, normal adjacent kidney tissues, and other cancer types. Our results revealed a major ccRCC molecular component correlating with cellular mitotic age-related features, particularly the mitotic epigenetic clock (age-adjusted epiTOC2), clock-like DNA mutational signatures (SBS1/ID1), and telomere attrition. This molecular component was overrepresented in ccRCC tumours in comparison with normal paired kidney tissues, associating with PBRM1 and SETD2 somatic cancer driver mutations, genome stability, tumor stage, grade and ccRCC patient survival, independently to chronological age. Pan-cancer analysis supported the mitotic-age effect, represented by this molecular component, in other cancer types. Another ccRCC component was associated with tobacco usage, the presence of tobacco related DNA mutational and methylation signatures, increased total mutation burden, and sex. This component was also related to the epigenetic regulation of xenobiotic metabolism-related genes (e.g., GSTP1), further suggesting a relationship with genotoxic compounds. We further identified molecular components related to the ccRCC tumour microenvironment and cell proliferation, including one component related to BAP1 cancer driver mutations, pro-inflammatory immune cells, and ccRCC patient survival. In conclusion, our study reports a novel framework to molecular characterize ccRCC tumours using an integrative multi-omic approach, providing additional insights into the molecular footprints of endogenous and exogenous exposures in ccRCC tumours, including molecular components with prognostic value for patients. Citation Format: Ricardo Cortez Cardoso Penha, Alexandra Sexton Oates, Sergey Senkin, Han La Park, Nicolas Alcala, Matthieu Foll, Karl Smith-Byrne, Paul Brennan, James Mckay. Multi-omic characterisation of clear cell renal carcinoma reveals endogenous and exogenous processes related to its aetiology [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6231.

OLIG2 is an in vivo bookmarking transcription factor in the developing neural tube in mouse.

Cells possess intrinsic features that are inheritable via epigenetic regulation, such as DNA methylation and histone modification. These inheritable features maintain a unique gene expression pattern, underlying cellular memory. Because of the degradation or displacement of mitotic chromosomes, most transcription factors do not contribute to cellular memory. However, accumulating in vitro evidence indicates that some transcription factors can be retained in mitotic chromosomes called as bookmarking. Such transcription factors may contribute to a novel third mechanism of cellular memory. Since most findings of transcription factor bookmarking have been reported in vitro, little is currently known in vivo. In the neural tube of mouse embryos, we discovered that OLIG2, a basic helix loop helix (bHLH) transcription factor that regulates proliferation of neural progenitors and the cell fate of motoneurons and oligodendrocytes, binds to chromatin through every cell cycle including M-phase. OLIG2 chromosomal localization coincides with mitotic cell features such as the phosphorylation of histone H3, KI67, and nuclear membrane breakdown. Chromosomal localization of OLIG2 is regulated by an N-terminus triple serine motif. Photobleaching analysis revealed slow OLIG2 mobility, suggesting a high affinity of OLIG2 to DNA. In Olig2 N-terminal deletion mutant mice, motoneurons and oligodendrocyte progenitor numbers are reduced in the neural tube, suggesting that the bookmarking regulatory domain is important for OLIG2 function. We conclude that OLIG2 is a de novo in vivo bookmarking transcription factor. Our results demonstrate the presence of in vivo bookmarking in a living organism and illustrate a novel function of transcription factors.

Histomorphological and Demographic Profile of Breast Carcinoma in Anatomical Pathology Department FKUI/RSCM 2016-2019

BackgroundInvasive breast carcinoma (IBC) is the most commonly diagnosed cancer in woman in the world and Indonesia. IBC is classified into two main categories: IBC of no special type (NST) and special types. This retrospective study aims to provide histomorphological profile of mastectomy cases of IBC in Anatomical Pathology Department, Faculty of Medicine, Universitas Indonesia, Dr. Cipto Mangunkusumo General Hospital (PA FKUI/RSCM) 2016-2019.MethodsThis is a descriptive research with cross-sectional design of mastectomy cases, using secondary data from the archives of PA FKUI/RSCM, starting from 2016- 2019. Collecting clinical data as well as observing several histopathologic parameters including histological type, grade, atypical mitotic features, lymphovascular invasion, and lymph node metastases. Tumour classification was made based on the WHO breast criteria in 2019.ResultsDuring the 2016-2019 period there were 144 cases of IBC. 99.3% of cases occurred in female patients, mostly in the age group from 40 to 59 years and unilateral. The majority had tumour sizes ranging >2-5 cm and were stage II. About a third of cases had received neoadjuvant chemotherapy, most commonly in advanced local stage tumors. Most histologic type (65.3%) were IBC of NST and 52.8% of cases were grade 2. There were 92 (63.4%) cases with atypical mitotic features, most commonly found in stage III. Lymphovascular invasion was found in 38.9% of cases and lymph node metastasis was found in 63.2% of cases.ConclusionIBC is the most common cancer in woman, with the most common histologic type were IBC, grade 2, stage II or more. Most of the cases show atypical mitosis, which is most commonly found in stage III. The findings of lymphovascular invasion did not always coincide with the findings of lymph node metastases.

CS-6 A case of poorly differentiated chordoma with systemic metastasis

A case report: The patient was a 32-year-old man with diplopia. He was diagnosed as sphenoid sinusitis on MRI by a local doctor and visited an otolaryngologist. MRI showed extensive extension of neoplastic lesions from the clivus to the sphenoid sinus to the anterior ethmoid sinuses, bilateral cavernous sinuses, and the right medial and lateral pterygoid muscles. The right Lebiere’s lymph node was enlarged and thought to be a metastatic site. Based on the rapid growth and extension of the tumor, the patient was referred to the Department of Otolaryngology at our hospital on suspicion of sinonasal carcinoma. The possibility of chordoma could not be denied, so the patient was referred to our department. The patient underwent a joint endoscopic extended transsphenoidal tumor resection. The pathological diagnosis showed mitotic and necrotic features, and the majority of the cells showed highly atypical components without mucous substrate. However, brachyury, a marker for chordoma, was diffusely positive, and there was loss of INI1 (SMARCB1) expression. The final diagnosis was poorly differentiated chordoma. Postoperatively, the tumor in the right cavernous sinus grew rapidly, and the right eye became blind due to obstruction of the superior ophthalmic vein. The patient was treated with Gamma Knife as soon as possible in the hope of local control by high-dose irradiation, and after a total of three irradiations, the residual tumor shrank markedly and symptoms improved, but systemic metastasis occurred in a short period of time and the patient died. The number of cases of poorly differentiated chordoma has been reported rarely (more than 50), and it is more common in children and even rarer in adults. We report this case with a review of the literature.

Protuberant Fibro-Osseous Lesion of the Temporal Bone: "Bullough Bump"-Multimodality Imaging Case Series and Literature Review.

A handful of cases of protuberant fibro-osseous lesions of the temporal bones have been described in the literature to date, with primary focus on the pathologic features. Here we review 3 cases of pathology-proved protuberant fibro-osseous lesions of the temporal bone and include a literature review with a focus on the imaging features. While rare, these lesions have near-pathognomonic imaging features defined by a location at the cortex of the outer table of the temporal bone at the occipitomastoid suture, lack of involvement of the underlying marrow, variable mineralization, and MR signal characteristics atypical of a chondroid lesion. One case in this series was FDG-avid and had occasional mitotic features, possibly reflecting an aggressive variant. Neuroradiologists should be familiar with this benign diagnosis to aid in timely identification and avoid unnecessary additional imaging.

CentTracker: a trainable, machine-learning-based tool for large-scale analyses of Caenorhabditis elegans germline stem cell mitosis.

Investigating the complex interactions between stem cells and their native environment requires an efficient means to image them in situ. Caenorhabditis elegans germline stem cells (GSCs) are distinctly accessible for intravital imaging; however, long-term image acquisition and analysis of dividing GSCs can be technically challenging. Here we present a systematic investigation into the technical factors impacting GSC physiology during live imaging and provide an optimized method for monitoring GSC mitosis under minimally disruptive conditions. We describe CentTracker, an automated and generalizable image analysis tool that uses machine learning to pair mitotic centrosomes and that can extract a variety of mitotic parameters rapidly from large-scale data sets. We employ CentTracker to assess a range of mitotic features in a large GSC data set. We observe spatial clustering of mitoses within the germline tissue but no evidence that subpopulations with distinct mitotic profiles exist within the stem cell pool. We further find biases in GSC spindle orientation relative to the germline’s distal–proximal axis and thus the niche. The technical and analytical tools provided herein pave the way for large-scale screening studies of multiple mitotic processes in GSCs dividing in situ, in an intact tissue, in a living animal, under seemingly physiological conditions.

Attention-Guided Multi-Branch Convolutional Neural Network for Mitosis Detection From Histopathological Images.

Mitotic count is an important indicator for assessing the invasiveness of breast cancers. Currently, the number of mitoses is manually counted by pathologists, which is both tedious and time-consuming. To address this situation, we propose a fast and accurate method to automatically detect mitosis from the histopathological images. The proposed method can automatically identify mitotic candidates from histological sections for mitosis screening. Specifically, our method exploits deep convolutional neural networks to extract high-level features of mitosis to detect mitotic candidates. Then, we use spatial attention modules to re-encode mitotic features, which allows the model to learn more efficient features. Finally, we use multi-branch classification subnets to screen the mitosis. Compared to existing related methods in literature, our method obtains the best detection results on the dataset of the International Pattern Recognition Conference (ICPR) 2012 Mitosis Detection Competition. Code has been made available at: https://github.com/liushaomin/MitosisDetection.

Fibrosarcoma pada anjing golden retriever di Rumah Sakit Hewan Pendidikan Universitas Airlangga

The dog named Agra (Golden Retriever) is suspected of having an axillary tumor. After surgery and examination of the pathology laboratory, it was found that the dog had fibrosarcoma in the axillaries. Fibrosarcoma is a malignant neoplasm that originates from mesenchymal cells, where the dominant cell histology is fibroblast cells that divide excessively and uncontrollably, can attack local tissues, and can move to other locations in the body (metastatic). The histopathological features of fibrosarcoma have fusiform or spindle-shaped cell fascicular growth patterns. The boundary between cells appears unclear with little cytoplasm and collagen fibers form parallel webbing. Grading histology is mainly based on the degree of cellularity, cell differentiation, mitotic features, and the amount of collagen produced by the cell necrosis. Surgery until now is the first and foremost thing in tumor therapy. Benign tumors can be removed with various surgical techniques depending on the location of the tumor. As for malignant tumors, surgery can be carried out by following other additional therapies to inhibit tumor growth.

A New Inhibitor of Tubulin Polymerization Kills Multiple Cancer Cell Types and Reveals p21-Mediated Mechanism Determining Cell Death after Mitotic Catastrophe.

Induction of mitotic catastrophe through the disruption of microtubules is an established target in cancer therapy. However, the molecular mechanisms determining the mitotic catastrophe and the following apoptotic or non-apoptotic cell death remain poorly understood. Moreover, many existing drugs targeting tubulin, such as vincristine, have reduced efficacy, resulting from poor solubility in physiological conditions. Here, we introduce a novel small molecule 2-aminoimidazoline derivative—OAT-449, a synthetic water-soluble tubulin inhibitor. OAT-449 in a concentration range from 6 to 30 nM causes cell death of eight different cancer cell lines in vitro, and significantly inhibits tumor development in such xenograft models as HT-29 (colorectal adenocarcinoma) and SK-N-MC (neuroepithelioma) in vivo. Mechanistic studies showed that OAT-449, like vincristine, inhibited tubulin polymerization and induced profound multi-nucleation and mitotic catastrophe in cancer cells. HeLa and HT-29 cells within 24 h of treatment arrested in G2/M cell cycle phase, presenting mitotic catastrophe features, and 24 h later died by non-apoptotic cell death. In HT-29 cells, both agents altered phosphorylation status of Cdk1 and of spindle assembly checkpoint proteins NuMa and Aurora B, while G2/M arrest and apoptosis blocking was consistent with p53-independent accumulation in the nucleus and largely in the cytoplasm of p21/waf1/cip1, a key determinant of cell fate programs. This is the first common mechanism for the two microtubule-dissociating agents, vincristine and OAT-449, determining the cell death pathway following mitotic catastrophe demonstrated in HT-29 cells.

Fundamental control of grade-specific colorectal cancer morphology by Src regulation of ezrin-centrosome engagement.

The phenotypic spectrum of colorectal cancer (CRC) is remarkably diverse, with seemingly endless variations in cell shape, mitotic figures and multicellular configurations. Despite this morphological complexity, histological grading of collective phenotype patterns provides robust prognostic stratification in CRC. Although mechanistic understanding is incomplete, previous studies have shown that the cortical protein ezrin controls diversification of cell shape, mitotic figure geometry and multicellular architecture, in 3D organotypic CRC cultures. Because ezrin is a substrate of Src tyrosine kinase that is frequently overexpressed in CRC, we investigated Src regulation of ezrin and morphogenic growth in 3D CRC cultures. Here we show that Src perturbations disrupt CRC epithelial spatial organisation. Aberrant Src activity suppresses formation of the cortical ezrin cap that anchors interphase centrosomes. In CRC cells with a normal centrosome number, these events lead to mitotic spindle misorientation, perturbation of cell cleavage, abnormal epithelial stratification, apical membrane misalignment, multilumen formation and evolution of cribriform multicellular morphology, a feature of low-grade cancer. In isogenic CRC cells with centrosome amplification, aberrant Src signalling promotes multipolar mitotic spindle formation, pleomorphism and morphological features of high-grade cancer. Translational studies in archival human CRC revealed associations between Src intensity, multipolar mitotic spindle frequency and high-grade cancer morphology. Collectively, our study reveals Src regulation of CRC morphogenic growth via ezrin-centrosome engagement and uncovers combined perturbations underlying transition to high-grade CRC morphology. © 2020 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

OSTEOSARCOMA OF THE JAWS: CLINICOPATHOLOGIC STUDY OF 3 CASES

Osteosarcoma is the most frequent primary malignant bone tumor, but osteosarcomas of the jaws are uncommon and account for only 2% to 10% of all cases diagnosed. The aim of this study is to present 3 new cases of osteosarcoma affecting the gnathic bones. The mandible was affected in 2 cases, whereas the maxilla was involved in 1. Age ranged from 10 to 25 years, and all patients were female. An asymptomatic expansive swelling was the chief complaint in all cases. Microscopically, a diffuse proliferation of pleomorphic cells with presence of osteoid material and variable number of atypical mitotic features were observed. Ki67 staining showed a proliferative index of approximately 15%, and positivity for D2-40 and CDK4 were also observed. All patients are currently under oncological treatment. Support: FAPEMIG. Osteosarcoma is the most frequent primary malignant bone tumor, but osteosarcomas of the jaws are uncommon and account for only 2% to 10% of all cases diagnosed. The aim of this study is to present 3 new cases of osteosarcoma affecting the gnathic bones. The mandible was affected in 2 cases, whereas the maxilla was involved in 1. Age ranged from 10 to 25 years, and all patients were female. An asymptomatic expansive swelling was the chief complaint in all cases. Microscopically, a diffuse proliferation of pleomorphic cells with presence of osteoid material and variable number of atypical mitotic features were observed. Ki67 staining showed a proliferative index of approximately 15%, and positivity for D2-40 and CDK4 were also observed. All patients are currently under oncological treatment. Support: FAPEMIG.

P178 SYNCHRONOUS TRIPLE CANCER OF THE ESOPHAGUS, GASTRIC CARDIA AND GIST OF THE STOMACH: A CASE REPORT AND REVIEW OF THE LITERATURE

Abstract Background A 82-year old man was admitted to our hospital in February 2019 for a thorough investigation to determine an incidential finding of a tumor of the stomach in a CT-Scan. He had no history of previous abdominal surgery or of serious illness. The initial symptoms were slight dysphagia, deterioriation in general condition and weight loss in the last months. Endoscopy of the stomach confirmed a poorly differentiated adenocarcinoma of the gastric cardia (intestinal type, Siewert Type II AEG) with the suspicion of paragastric lymph node metastasis. The consensus of our Comprehensive Cancer Center was a neoadjuvant FLOT-chemotherapy followed by radical resection. The Patient presented again during chemotherapy in April 2019 complaining of discomfort and epigastric pain. Endoscopy showed a partial regress of the primary tumor and on CT scan a progress of the potential lymph node metastases. Due to the symptoms and findings early resection was recommended. On surgery extensive tumor masses were found around the lesser curve and around the peritoneal cavity not like lymph node metastases. An radical resection with intrathoracic anastomoses with gastric pull up was performed. The patient was discharged 4 weeks postoperatively. The histopathological findings showed 2 other synchronous cancers besides the adenocarcinoma of the gastric cardia (ypT1b, L0;V1,G3,R0, HER2-negative, no microsatellite instability). It showed an early stage of squamous cell carcinoma of the esophagus (pT1a (m3-Type), Gn0 (0/21), L0, V0, G1, R0) and the potential paragastric lymphnode metastasis revealed to be a rare high-risk wild-type GIST of the stomach which showed a high mitotic index (> 120 mitoses/15 high-power fields). The patient was readmitted to our hospital because of severe epigastric pain. A CT-Scan showed a massive progress and local recurrence of the GIST with a diameter of 7cm and infiltration into the portal vein and ascites with suspicion on peritoneal carcinomatosis. Despite the attempt of a sunitinib therapy the patient died in the beginning of May 2019 in a palliative setting . Conclusion There have only been few reports in the literature of synchronous triple cancer of gastric GIST, esophageal squamous cell carcinoma, and gastric adenocarcinoma. In our case we considered the gastric adenocarcinoma to be a middle stage cancer and the squamous cell carcinoma an early stage esophageal cancer. The incidental finding of the high-grade gastric GIST with a high mitotic index and unfavorable histopathological features as the wild-type confirmation was probably the cause of death of the patient.

Implementation of various approaches to study the prevalence, incidence and progression of disseminated neoplasia in mussel stocks

Marine mussel production is of substantial economic interest in numerous coastal areas worldwide, making crucial the study of pathologies that affect them. Disseminated neoplasia (DN) has recently been suggested to be linked to blue mussel, Mytilus edulis, mortality outbreaks observed in France since 2014, although the evidence remains indirect. In order to improve DN detection and monitoring, we compared the sensitivity of four diagnostic tools, namely haemocytology, histology, flow cytometry, and genetics. Haemocytological examination gave the best results in sensitivity and had the advantage of being non-invasive, allowing disease progression to be followed in affected mussels. Using this approach, we showed that DN progression is usually slow, and we provide evidence of remission events. We observed a high diversity of forms and mitotic features of neoplastic cells located in the vesicular connective tissue but rarely in the haemolymph. Circulating cells occur as four main types but are homogenous in morphology and DNA content within a single individual. Polyploidy proved very high, from 8 N to 18 N. Genetic analysis of haemolymph DNA showed that a Mytilus trossulus genetic signal was associated with almost all the DN cases here diagnosed by haemocytological examination, regardless of the DN type. This result corroborates DN is a transmissible cancer that first originated in a M. trossulus host and subsequently crossed into M. edulis. No pre-neoplastic conditions were detectable. The prevalence of the disease was quite low, which, together with the low morbidity observed in the lab, suggest DN is unlikely to be the direct cause of mortality outbreaks in France.

Invasive stratified mucin-producing carcinoma: a clinicopathological analysis of three cases

ABSTRACTObjective: To investigate the clinicopathologic features and immunophenotype of invasive-stratified mucin-producing carcinoma (ISMC).Methods: We retrospectively analyzed three patients who underwent surgery for pathologically confirmed ISMC from January 2017 to December 2017 in Anyang Tumor Hospital. The pathological features, immunophenotype, and prognosis were discussed.Results: Clinical symptoms and imaging were atypical. Microscopically, the arrangement of tumor cells is close to that of squamous cell carcinoma, showing flake or nested growth. Morphologically, tumor cells were reminiscent of adenocarcinoma. The tumor cells showed abundant intracytoplasmic mucus which routinely created spacing between adjacent nuclei. The nuclei was round or oval with irregular karyotypes, thick nuclear membrane, staining empty light, rough chromatin, visible small nucleoli. Mucous vacuoles or pink-stained foam were visible in the cytoplasm. Mitotic features and apoptotic bodies were seen in the invasive component of all three cases, whereas all but one of these showed a neutrophil-dominant inflammatory cells infiltration. A large number of histocytes were found in the stroma of two cases. Special staining of mucus confirmed the presence of mucus in cytoplasm. Immunohistochemical results showed that tumor cells were strongly and diffusely positive for p16 and CK7 and had high Ki-67 expression.Conclusions: ISMC is a rare tumor in female genital tract and is often misdiagnosed as squamous cell carcinoma and other cervical cancers. The diagnosis mainly depends on the unique clinicopathologic features together with the immunophenotype. Treatment and prognosis, like other cervical cancer, are mainly based on clinical stages.

Abstract P3-08-08: SOX11 is a potential prognostic marker of high-risk breast ductal carcinoma in situ

Abstract Ductal carcinoma in situ (DCIS) comprises 20-25% of screen-detected breast cancers and, like invasive ductal carcinoma (IDC), is heterogenous in terms of the underlying biology, presentation, and outcome. While there are limited potential biomarkers of outcome for DCIS, estrogen receptor (ER)- positive, progesterone receptor (PR)- positive, and HER2- negative DCIS appears to have a better prognosis compared to ER- negative, PR- negative, and HER2- positive DCIS. The aim of this study was to identify additional clinically relevant markers to stratify DCIS according to risk of relapse or progression to invasive disease. In order to determine the driver genes involved in DCIS evolution, we utilized transcriptional data sets (GSE788, GSE16873), containing data from both normal mammary glands (NMG) and DCIS. Upon performing class comparison (NMG vs DCIS), we identified 297 over-expressed genes and 187 under-expressed genes. The over-expressed genes represented mitotic and proliferative features annotated as mitotic spindle and condensed chromosomes, while the under-expressed genes were associated with loss of epithelial features annotated as epithelial cell differentiation and development. The 484 differentially expressed genes were further correlated with recurrence events using Kessler's breast cancer data set to identify genes contributing to the aggressive features across IDC and subsequently associated with DCIS. Genes correlating with recurrence events were selected. Of the 484 genes, 99 genes were found to be significantly associated with recurrence events of IDC (with P<0.003). Among these 99 genes, component genes of the Oncotype DCIS score and genes reported as relevant to DCIS biology were included for Nanostring transcriptomic analysis. The final number of genes-of-interest were 58, including 5 housekeeping genes. 40 DCIS lesions and 8 NMG tissue were macro- dissected from formalin- fix paraffin- embedded blocks (FFPE) and extracted transcripts were subjected for Nanostring analysis. Gene expression data was clustered in an unsupervised manner using R software. Two sample clusters were identified: an ER/PR- negative cluster and an ER/PR- positive cluster. Over-expression of transcription factor SOX11, along with HER2, was exclusively seen in the ER/PR- negative cluster. This cluster was further categorized into HER2-low/SOX11+ and HER2-high/SOX11+ groups. These RNA expression findings are undergoing confirmation by immunohistochemistry (IHC) of the FFPE tumor sections. An independent series of 15 DCIS cases that have recurred as DCIS or progressed to IDC were analyzed by IHC, revealing SOX11 expression only present in cases displaying a high proportion of HER2+ expression. SOX11 is exclusively expressed in ER/PR-negative DCIS and is a candidate clinical marker for recurrence of DCIS or progression to IDC. Citation Format: Ladha FA, Kai K, Lu W, Yang F, Tang X, Wistuba II, Sen S, Thompson A. SOX11 is a potential prognostic marker of high-risk breast ductal carcinoma in situ [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P3-08-08.

Dinoflagellate nucleus contains an extensive endomembrane network, the nuclear net

Dinoflagellates are some of the most common eukaryotic cells in the ocean, but have very unusual nuclei. Many exhibit a form of closed mitosis (dinomitosis) wherein the nuclear envelope (NE) invaginates to form one or more trans-nuclear tunnels. Rather than contact spindles directly, the chromatids then bind to membrane-based kinetochores on the NE. To better understand these unique mitotic features, we reconstructed the nuclear architecture of Polykrikos kofoidii in 3D using focused ion beam scanning electron microscopy (FIB-SEM) in conjunction with high-pressure freezing, freeze-substitution, TEM, and confocal microscopy. We found that P. kofoidii possessed six nuclear tunnels, which were continuous with a reticulating network of membranes that has thus far gone unnoticed. These membranous extensions interconnect the six tunnels while ramifying throughout the nucleus to form a “nuclear net.” To our knowledge, the nuclear net is the most elaborate endomembrane structure described within a nucleus. Our findings demonstrate the utility of tomographic approaches for detecting 3D membrane networks and show that nuclear complexity has been underestimated in Polykrikos kofoidii and, potentially, in other dinoflagellates.

The Effect of Zinc-Deficient Pregnant Albino Rats on the Retinal Development of their Embryos and Offspring: Light Microscopic and Immunohistochemical Study

AbstractBackground: Zinc is involved in many biochemical func-tions. Over 300 enzymes require zinc for their activation and nearly 2000 transcription factors require zinc for gene expres-sion. Zinc is also an effective antioxidant and anti-inflammatory agent. Nutritional deficiency of zinc may affect nearly 2 billion subjects in the developing world.Aim of Study: Because of the prevalence of malnutrition throughout the world this study was carried out to detect the possible effects of zinc-deficient pregnant albino rats on the retinal development of their embryos and offspring.Material and Methods: Female rats after mating were randomly divided into 2 equal groups: One control group received single intraperitoneal injection of distilled water on day 9 of gestation and one experimental group received intraperitoneal injections of 1.10 phenanthroline (zinc chelator purchased from Sigma Company), in a single dose of 30 mg/kg on day 9 of gestation. From the embryos and offspring of both control and treated mothers (at the following prenatal gestational days: 13, 15, 17, 19 and the following postnatal days: (newborn, 7 days, 15 days and 21 days), retinal specimens were processed for Haematoxylin & Eosin staining technique and immunohistochemical study using caspase-3 antibody was used to detect apoptosis in the postnatal age groups.Results: The light microscopic study of the 13 days zinc deficient embryo retina showed apparent shrinkage of the lens vesicle and the optic cup was well formed but it appeared to be shrunken. The retinal pigment epithelium (RPE) of 15th, 17th and 19th day old zinc-deficient embryos consistently contained apoptotic cells with vacuolated cytoplasm and rarified nuclei. The neuroblastic layer showed apparent de-crease in thickness and less mitotic features. The ganglion cell layer revealed numerous deeply stained pyknotic nuclei in the 19th gestational day. Retinae of the newborn, 7 days, 15 days, 21 days-old zinc deficient albino rats showed that the (RPE) contained some vacuolated cells with pyknotic nuclei. The organization of the retina was altered in zinc deficient rats. In addition, apparent reduction in the total retinal thickness and degenerative changes in some layers were observed. By the immunohistochemical study, there was intense positive caspase-3 reaction in all layers of the treated rat retina.Conclusion: This study revealed that zinc has a role in the developing rat retina and its deficiency causes cytoarchi-tectural and maturational changes in the retina of the albino rats. These histological changes may impair the visual functions of the retina; so it is recommended that further investigations should be carried out to determine the relevancy of deprivation of zinc as an etiologic factor in the development of retinal degeneration.

Incidence and clinical outcomes of atypical pituitary adenomas

BackgroundAtypical pituitary adenoma has a high rate of recurrence and most of them need further treatment after surgery. This is why early detection is very important.MethodsIn this study we examined the incidence and outcome of large atypical pituitary adenoma in our patient cohort. The study was a retrospective review of fifty consecutive patients at Assiut university hospital. They all had a large adenoma with supracellar and parasellar extension. They were considered unsuitable for transphenoidal approach. They all had craniotomy. This study was carried out during a thirty eight month period. Pathology reports were reviewed. All patients had MIB-1 immunohistochemical data. Patients with lesions showing excessive MIB-1immunoreactivity and mitotic features had p53 immunohistochemical data.ResultsTen (20%) of our patients had lesions that met all the criteria for atypical adenoma.ConclusionAtypical pituitary adenoma tend to be aggressive and invasive macroadenomas. Following surgery, further treatment such as radiotherapy should be considered.

Ultrastructural analysis of mitotic Drosophila S2 cells identifies distinctive microtubule and intracellular membrane behaviors

BackgroundS2 cells are one of the most widely used Drosophila melanogaster cell lines. A series of studies has shown that they are particularly suitable for RNAi-based screens aimed at the dissection of cellular pathways, including those controlling cell shape and motility, cell metabolism, and host–pathogen interactions. In addition, RNAi in S2 cells has been successfully used to identify many new mitotic genes that are conserved in the higher eukaryotes, and for the analysis of several aspects of the mitotic process. However, no detailed and complete description of S2 cell mitosis at the ultrastructural level has been done. Here, we provide a detailed characterization of all phases of S2 cell mitosis visualized by transmission electron microscopy (TEM).ResultsWe analyzed by TEM a random sample of 144 cells undergoing mitosis, focusing on intracellular membrane and microtubule (MT) behaviors. This unbiased approach provided a comprehensive ultrastructural view of the dividing cells, and allowed us to discover that S2 cells exhibit a previously uncharacterized behavior of intracellular membranes, involving the formation of a quadruple nuclear membrane in early prometaphase and its disassembly during late prometaphase. After nuclear envelope disassembly, the mitotic apparatus becomes encased by a discontinuous network of endoplasmic reticulum membranes, which associate with mitochondria, presumably to prevent their diffusion into the spindle area. We also observed a peculiar metaphase spindle organization. We found that kinetochores with attached k-fibers are almost invariably associated with lateral MT bundles that can be either interpolar bundles or k-fibers connected to a different kinetochore. This spindle organization is likely to favor chromosome alignment at metaphase and subsequent segregation during anaphase.ConclusionsWe discovered several previously unknown features of membrane and MT organization during S2 cell mitosis. The genetic determinants of these mitotic features can now be investigated, for instance by using an RNAi-based approach, which is particularly easy and efficient in S2 cells.

Fine-Tuned and Cell-Cycle-Restricted Expression of Fusogenic Protein Syncytin-2 Maintains Functional Placental Syncytia

Many types of multinucleated cells (syncytia) generated by cell-cell fusion are post-mitotic, but it remains unclear how this state is maintained and why. Here, we utilized the fluorescent ubiquitination-based cell-cycle indicator (Fucci) reporter system to show that human placental trophoblast cells were all in the G0 phase before they fuse. Expression of the fusogenic protein (fusogen) Syncytin-2 was confined to G0 cells. Overexpression of Syncytin-2 in cycling cells overrode the cell-cycle restriction and enabled fusion of cells in the S/G2/M phases but resulted in the unstable syncytia retaining mitotic features. The Syncytin-2-induced syncytia were functionally compromised with respect to pathogen defense and hormone secretion. We found that, during trophoblast fusion, the cell-cycle inhibitor p21 interacted with the GCM1 transcription factor, and this complex bound to the promoter of Syncytin-2 and promoted its transcription. These findings demonstrate that G0-restricted Syncytin-2 expression is aprerequisite for development of functional post-mitotic syncytia.