Pathology - Research Publications
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ItemTight Junction Protein Claudin-2 Promotes Self-Renewal of Human Colorectal Cancer Stem-like Cells(American Association for Cancer Research, 2018-06-01)Posttreatment recurrence of colorectal cancer, the third most lethal cancer worldwide, is often driven by a subpopulation of cancer stem cells (CSC). The tight junction (TJ) protein claudin-2 is overexpressed in human colorectal cancer, where it enhances cell proliferation, colony formation, and chemoresistance in vitro. While several of these biological processes are features of the CSC phenotype, a role for claudin-2 in the regulation of these has not been identified. Here, we report that elevated claudin-2 expression in stage II/III colorectal tumors is associated with poor recurrence-free survival following 5-fluorouracil–based chemotherapy, an outcome in which CSCs play an instrumental role. In patient-derived organoids, primary cells, and cell lines, claudin-2 promoted colorectal cancer self-renewal in vitro and in multiple mouse xenograft models. Claudin-2 enhanced self-renewal of ALDHHigh CSCs and increased their proportion in colorectal cancer cell populations, limiting their differentiation and promoting the phenotypic transition of non-CSCs toward the ALDHHigh phenotype. Next-generation sequencing in ALDHHigh cells revealed that claudin-2 regulated expression of nine miRNAs known to control stem cell signaling. Among these, miR-222-3p was instrumental for the regulation of self-renewal by claudin-2, and enhancement of this self-renewal required activation of YAP, most likely upstream from miR-222-3p. Taken together, our results indicate that overexpression of claudin-2 promotes self-renewal within colorectal cancer stem-like cells, suggesting a potential role for this protein as a therapeutic target in colorectal cancer.
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ItemNo Preview AvailableFailure of Autophagy-Lysosomal Pathways in Rod Photoreceptors Causes the Early Retinal Degeneration Phenotype Observed in Cln6(nclf) Mice(ASSOC RESEARCH VISION OPHTHALMOLOGY INC, 2018-10-01)Purpose: Vision loss caused by photoreceptor death represents one of the first symptoms in neuronal ceroid lipofuscinosis, a condition characterized by accumulation of intracellular waste. Cln6nclf mice have a naturally occurring mutation in ceroid-lipofuscinosis neuronal (CLN) protein 6 and are a model of this disorder. In order to identify the effect intracellular waste (lipofuscin) accumulation plays in driving retinal degeneration, the time course of degeneration was carefully characterized functionally using the electroretinogram and structurally using histology. Methods: Cln6nclf and C57BL/6J, wild-type, mice were studied at postnatal day 18 (P18), P30, P60, P120, and P240, and retinal degeneration was correlated with changes in the retinal pigment epithelial (RPE) and neuronal autophagy-lysosomal pathways using super-resolution microscopy. Results: In Cln6nclf mice there was significant loss of rod photoreceptor function at P18, prior to photoreceptor nuclei loss at P60. In contrast, cone pathway function was not affected until P240. The loss of rod photoreceptor function correlated with significant disruption of the autophagy-lysosomal degradation pathways within photoreceptors, but not in the RPE or other retinal neurons. Additionally, there was cytosolic accumulation of P62 and undigested mitochondrial-derived, ATP synthase subunit C in the photoreceptor layers of Cln6nclf mice at P30. Conclusions: These results suggest that rod photoreceptors have an increased sensitivity to disturbances in the autophagy-lysosomal pathway and the subsequent failure of mitochondrial turnover, relative to other retinal cells. It is likely that primary failure of the rod photoreceptors rather than the RPE or other retinal neurons underlies the early visual dysfunction that occurs in the Cln6nclf mouse model.
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ItemPhosphorylation of Acetyl-CoA Carboxylase by AMPK Reduces Renal Fibrosis and Is Essential for the Anti-Fibrotic Effect of Metformin(AMER SOC NEPHROLOGY, 2018-09-01)BACKGROUND: Expression of genes regulating fatty acid metabolism is reduced in tubular epithelial cells from kidneys with tubulointerstitial fibrosis (TIF), thus decreasing the energy produced by fatty acid oxidation (FAO). Acetyl-CoA carboxylase (ACC), a target for the energy-sensing AMP-activating protein kinase (AMPK), is the major controller of the rate of FAO within cells. Metformin has a well described antifibrotic effect, and increases phosphorylation of ACC by AMPK, thereby increasing FAO. METHODS: We evaluated phosphorylation of ACC in cell and mouse nephropathy models, as well as the effects of metformin administration in mice with and without mutations that reduce ACC phosphorylation. RESULTS: Reduced phosphorylation of ACC on the AMPK site Ser79 occurred in both tubular epithelial cells treated with folate to mimic cellular injury and in wild-type (WT) mice after induction of the folic acid nephropathy model. When this effect was exaggerated in mice with knock-in (KI) Ser to Ala mutations of the phosphorylation sites in ACC, lipid accumulation and fibrosis increased significantly compared with WT. The effect of ACC phosphorylation on fibrosis was confirmed in the unilateral ureteric obstruction model, which showed significantly increased lipid accumulation and fibrosis in the KI mice. Metformin use was associated with significantly reduced fibrosis and lipid accumulation in WT mice. In contrast, in the KI mice, the drug was associated with worsened fibrosis. CONCLUSIONS: These data indicate that reduced phosphorylation of ACC after renal injury contributes to the development of TIF, and that phosphorylation of ACC is required for metformin's antifibrotic action in the kidney.
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ItemAmyloid precursor protein and amyloid precursor-like protein 2 have distinct roles in modulating myelination, demyelination, and remyelination of axons(WILEY, 2019-03-01)The identification of factors that regulate myelination provides important insight into the molecular mechanisms that coordinate nervous system development and myelin regeneration after injury. In this study, we investigated the role of amyloid precursor protein (APP) and its paralogue amyloid precursor-like protein 2 (APLP2) in myelination using APP and APLP2 knockout (KO) mice. Given that BACE1 regulates myelination and myelin sheath thickness in both the peripheral and central nervous systems, we sought to determine if APP and APLP2, as alternate BACE1 substrates, also modulate myelination, and therefore provide a better understanding of the events regulating axonal myelination. In the peripheral nervous system, we identified that adult, but not juvenile KO mice, have lower densities of myelinated axons in their sciatic nerves while in the central nervous system, axons within both the optic nerves and corpus callosum of both KO mice were significantly hypomyelinated compared to wild-type (WT) controls. Biochemical analysis demonstrated significant increases in BACE1 and myelin oligodendrocyte glycoprotein and decreased NRG1 and proteolipid protein levels in both KO brain tissue. The acute cuprizone model of demyelination/remyelination revealed that whereas axons in the corpus callosum of WT and APLP2-KO mice underwent similar degrees of demyelination and subsequent remyelination, the myelinated callosal axons in APP-KO mice were less susceptible to cuprizone-induced demyelination and showed a failure in remyelination after cuprizone withdrawal. These data identified APP and APLP2 as modulators of normal myelination and demyelination/remyelination conditions. Deletion of APP and APLP2 identifies novel interplays between the BACE1 substrates in the regulation of myelination.
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ItemSurgical stress response and promotion of metastasis in colorectal cancer: a complex and heterogeneous process(SPRINGER, 2018-04-01)Surgery remains the curative treatment modality for colorectal cancer in all stages, including stage IV with resectable liver metastasis. There is emerging evidence that the stress response caused by surgery as well as other perioperative therapies such as anesthesia and analgesia may promote growth of pre-existing micro-metastasis or potentially initiate tumor dissemination. Therapeutically targeting the perioperative period may therefore reduce the effect that surgical treatments have in promoting metastases, for example by combining β-adrenergic receptor antagonists and cyclooxygenase-2 (COX-2) inhibitors in the perioperative setting. In this paper, we highlight some of the mechanisms that may underlie surgery-related metastatic development in colorectal cancer. These include direct tumor spillage at the time of surgery, suppression of the anti-tumor immune response, direct stimulatory effects on tumor cells, and activation of the coagulation system. We summarize in more detail results that support a role for catecholamines as major drivers of the pro-metastatic effect induced by the surgical stress response, predominantly through activation of β-adrenergic signaling. Additionally, we argue that an improved understanding of surgical stress-induced dissemination, and more specifically whether it impacts on the level and nature of heterogeneity within residual tumor cells, would contribute to the successful clinical targeting of this process. Finally, we provide a proof-of-concept demonstration that ex-vivo analyses of colorectal cancer patient-derived samples using RGB-labeling technology can provide important insights into the heterogeneous sensitivity of tumor cells to stress signals. This suggests that intra-tumor heterogeneity is likely to influence the efficacy of perioperative β-adrenergic receptor and COX-2 inhibition, and that ex-vivo characterization of heterogeneous stress response in tumor samples can synergize with other models to optimize perioperative treatments and further improve outcome in colorectal and other solid cancers.
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ItemPre-eclampsia is associated with altered expression of the renal sodium transporters NKCC2, NCC and ENaC in urinary extracellular vesicles(PUBLIC LIBRARY SCIENCE, 2018-09-24)Pre-eclampsia is a hypertensive disorder of pregnancy characterised by hypertension and sodium retention by the kidneys. To identify changes in sodium uptake proteins in the tubules of the distal nephron, we studied their expression in urinary extracellular vesicles or exosomes (uEVs). Urine was collected from women with pre-eclampsia or during normal pregnancy, and from healthy non-pregnant controls. uEVs were isolated by centrifugation and analyzed by Western blot. Expression, proteolytic cleavage and phosphorylation was determined by densitometric analysis normalized to the exosome marker CD9. Results showed a significant increase in phosphorylation of the activating S130 site in NKCC2, the drug target for frusemide, in women with pre-eclampsia compared with normal pregnant women. Phosphorylation of the activating sites T101/105 in NKCC2 was similar but the activating T60 site in NCC, the drug target for thiazide diuretics, showed significantly less phosphorylation in pre-eclampsia compared with normal pregnancy. Expression of the larger forms of the α subunit of ENaC, the drug target for amiloride, was significantly greater in pre-eclampsia, with more fragmentation of theγ subunit. The differences observed are predicted to increase the activity of NKCC2 and ENaC while reducing that of NCC. This will increase sodium reabsorption, and so contribute to hypertension in pre-eclampsia.
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ItemOn the Developmental Timing of Stress: Delineating Sex-Specific Effects of Stress across Development on Adult Behavior(MDPI, 2018-07-01)Stress, and the chronic overactivation of major stress hormones, is associated with several neuropsychiatric disorders. However, clinical literature on the exact role of stress either as a causative, triggering, or modulatory factor to mental illness remains unclear. We suggest that the impact of stress on the brain and behavior is heavily dependent on the developmental timing at which the stress has occurred, and as such, this may contribute to the overall variability reported on the association of stress and mental illness. Here, animal models provide a way to comprehensively assess the temporal impact of stress on behavior in a controlled manner. This review particularly focuses on the long-term impact of stress on behavior in various rodent stress models at three major developmental time points: early life, adolescence, and adulthood. We characterize the various stressor paradigms into physical, social, and pharmacological, and discuss commonalities and differences observed across these various stress-inducing methods. In addition, we discuss here how sex can influence the impact of stress at various developmental time points. We conclude here that early postnatal life and adolescence represent particular periods of vulnerability, but that stress exposure during early life can sometimes lead to resilience, particularly to fear-potentiated memories. In the adult brain, while shorter periods of stress tended to enhance spatial memory, longer periods caused impairments. Overall, males tended to be more vulnerable to the long-term effects of early life and adolescent stress, albeit very few studies incorporate both sexes, and further well-powered sex comparisons are needed.
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ItemAxillary lymphoma masquerading as inflammatory breast cancer.(Department of Biomedical Imaging, University of Malaya, Malaysia, 2006-07)Primary non-Hodgkins lymphoma (NHL) of the breast, and its extranodal spread to the breast resulting from systemic lymphoma, are recognised albeit uncommon conditions. However, lymphoma involving the axilla, presenting with the clinical appearance of inflammatory breast carcinoma (IBC) without infiltration of breast dermal lymphatics has not been reported previously.As highlighted by the two cases presented here, this entity should be considered in the differential diagnosis of patients presenting with clinical IBC. The cases highlight the importance of careful histological analysis to distinguish IBC from NHL, since management strategies and prognosis are quite different.
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ItemNo Preview AvailableFamilial occurrence of warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome(SPRINGER BASEL AG, 2008-12-01)INTRODUCTION: Warts, hypogammaglobulinemia, infections, and myelokathexis (WHIM) syndrome is a rare immunodeficiency disorder with an autosomal-dominant pattern of inheritance and low fatality rate but significant lifelong morbidity. MATERIALS AND METHODS: A 27-year-old mother of two children has been suffering from severe neutropenia and recurrent infections with the diagnosis of sporadic WHIM syndrome established by sequencing the CXCR4 gene and the finding of a heterozygous 1000 C-->T nonsense mutation in the second CXCR4 exon. The first child was an apparently healthy boy delivered at full term. Umbilical cord blood cells were obtained for genetic analysis. Peripheral blood cells were also analyzed at 8 months of life. Both analyses revealed the same mutation as that of his mother. The child was in a good condition, manifesting neutropenia without infections until 11 months of life. He subsequently developed pneumonia requiring a more aggressive treatment. After that, the regular substitution of immunoglobulins (IVIGs) and G-CSF has been preventing serious infections. Six months ago the second boy was delivered who also demonstrated neutropenia without severe infections. Genetic studies using cord blood and also peripheral blood cells in the fourth month showed an identical mutation of the CXCR4 gene as in his mother. Moreover, the mother and her first son demonstrated monocytopenia. RESULTS: The results indicate that genetic defects connected with WHIM syndrome may influence not only the granulocyte, but also the monocytic lineage. Moreover, a perinatal diagnosis of WHIM syndrome made by sequencing the CXCR4 gene should be performed in cases where either parent is known to be affected with this disease. CONCLUSIONS: This would facilitate an earlier detection of the deficiency in children, thereby allowing a more comprehensive follow-up and administration of appropriate therapy.
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ItemNo Preview AvailableMolecular characteristics of screen-detected vs symptomatic breast cancers and their impact on survival(NATURE PUBLISHING GROUP, 2009-10-13)BACKGROUND: Several recent studies have shown that screen detection remains an independent prognostic factor after adjusting for disease stage at presentation. This study compares the molecular characteristics of screen-detected with symptomatic breast cancers to identify if differences in tumour biology may explain some of the survival benefit conferred by screen detection. METHODS: A total of 1379 women (aged 50-70 years) with invasive breast cancer from a large population-based case-control study were included in the analysis. Individual patient data included tumour size, grade, lymph node status, adjuvant therapy, mammographic screening status and mortality. Immunohistochemistry was performed on tumour samples using 11 primary antibodies to define five molecular subtypes. The effect of screen detection compared with symptomatic diagnosis on survival was estimated after adjustment for grade, nodal status, Nottingham Prognostic Index (NPI) and the molecular markers. RESULTS: Fifty-six per cent of the survival benefit associated with screen-detected breast cancer was accounted for by a shift in the NPI, a further 3-10% was explained by the biological variables and more than 30% of the effect remained unexplained. CONCLUSION: Currently known biomarkers remain limited in their ability to explain the heterogeneity of breast cancer fully. A more complete understanding of the biological profile of breast tumours will be necessary to assess the true impact of tumour biology on the improvement in survival seen with screen detection.