RESUMO
Objective and Impact Statement: Human epidermal growth factor receptor 2 (HER2) is a critical protein in cancer cell growth that signifies the aggressiveness of breast cancer (BC) and helps predict its prognosis. Here, we introduce a deep learning-based approach utilizing pyramid sampling for the automated classification of HER2 status in immunohistochemically (IHC) stained BC tissue images. Introduction: Accurate assessment of IHC-stained tissue slides for HER2 expression levels is essential for both treatment guidance and understanding of cancer mechanisms. Nevertheless, the traditional workflow of manual examination by board-certified pathologists encounters challenges, including inter- and intra-observer inconsistency and extended turnaround times. Methods: Our deep learning-based method analyzes morphological features at various spatial scales, efficiently managing the computational load and facilitating a detailed examination of cellular and larger-scale tissue-level details. Results: This approach addresses the tissue heterogeneity of HER2 expression by providing a comprehensive view, leading to a blind testing classification accuracy of 84.70%, on a dataset of 523 core images from tissue microarrays. Conclusion: This automated system, proving reliable as an adjunct pathology tool, has the potential to enhance diagnostic precision and evaluation speed, and might substantially impact cancer treatment planning.
RESUMO
D-Serine is a physiological co-agonist that activates N-methyl D-aspartate receptors (NMDARs) and is essential for neurotransmission, synaptic plasticity, and behavior. D-Serine may also trigger NMDAR-mediated neurotoxicity, and its dysregulation may play a role in neurodegeneration. D-Serine is synthesized by the enzyme serine racemase (SR), which directly converts L-serine to D-serine. However, many aspects concerning the regulation of D-serine production under physiological and pathological conditions remain to be elucidated. Here, we investigate possible mechanisms regulating the synthesis of D-serine by SR in paradigms relevant to neurotoxicity. We report that SR undergoes nucleocytoplasmic shuttling and that this process is dysregulated by several insults leading to neuronal death, typically by apoptotic stimuli. Cell death induction promotes nuclear accumulation of SR, in parallel with the nuclear translocation of GAPDH and Siah proteins at an early stage of the cell death process. Mutations in putative SR nuclear export signals (NESs) elicit SR nuclear accumulation and its depletion from the cytosol. Following apoptotic insult, SR associates with nuclear GAPDH along with other nuclear components, and this is accompanied by complete inactivation of the enzyme. As a result, extracellular D-serine concentration is reduced, even though extracellular glutamate concentration increases severalfold. Our observations imply that nuclear translocation of SR provides a fail-safe mechanism to prevent or limit secondary NMDAR-mediated toxicity in nearby synapses.
Assuntos
Núcleo Celular/enzimologia , Neurônios/enzimologia , Racemases e Epimerases/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Serina/biossíntese , Sinapses/metabolismo , Transporte Ativo do Núcleo Celular/genética , Animais , Núcleo Celular/genética , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/genética , Gliceraldeído-3-Fosfato Desidrogenase (Fosforiladora)/metabolismo , Camundongos , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Racemases e Epimerases/genética , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/genética , Serina/genética , Sinapses/genéticaRESUMO
D-Serine and glycine are coagonists of NMDA receptors (NMDARs), but their relative contributions for several NMDAR-dependent processes are unclear. We now report that the alanine-serine-cysteine transporter-1 (Asc-1) mediates release of both D-serine and glycine from neurons, and, in turn, this modulates NMDAR synaptic activity. Asc-1 antiporter activity is enhanced by D-isoleucine (D-Ile), which releases D-serine and glycine from Asc-1-transfected cells, primary neuronal cultures, and hippocampal slices. D-Ile has no effect on astrocytes, which do not express Asc-1. We show that D-Ile enhances the long-term potentiation (LTP) in rat and mouse hippocampal CA1 by stimulating Asc-1-mediated endogenous D-serine release. D-Ile effects on synaptic plasticity are abolished by enzymatically depleting D-serine or by using serine racemase knock-out (SR-KO) mice, confirming its specificity and supporting the notion that LTP depends mostly on D-serine release. Conversely, our data also disclose a role of glycine in activating synaptic NMDARs. Although acute enzymatic depletion of D-serine also drastically decreases the isolated NMDAR synaptic potentials, these responses are still enhanced by D-Ile. Furthermore, NMDAR synaptic potentials are preserved in SR-KO mice and are also enhanced by D-Ile, indicating that glycine overlaps with D-serine binding at synaptic NMDARs. Altogether, our results disclose a novel role of Asc-1 in regulating NMDAR-dependent synaptic activity by mediating concurrent non-vesicular release of D-serine and glycine. Our data also highlight an important role of neuron-derived D-serine and glycine, indicating that astrocytic D-serine is not solely responsible for activating synaptic NMDARs.
