All-in-focus fine needle aspiration biopsy imaging based on Fourier ptychographic microscopy

Context: Cytology is the study of whole cells in diagnostic pathology. Unlike standard histologic thinly sliced specimens, cytologic preparations consist of preparations of whole cells where cells commonly cluster and aggregate. As such, cytology preparations are generally much thicker than histologic slides, resulting in large patches of defocus when examined under the microscope. A diagnostic aggregate of cells often cannot be viewed in focus together, requiring pathologists to continually manipulate the focal plane, complicating the task of accurately assessing the entire cellular aggregate and thus in making a diagnosis. Further, it is extremely difficult to acquire useful uniformly in-focus digital images of cytology preparations for applications such as remote diagnostic evaluations and artificial intelligence models. The predominant current method to address this issue is to acquire digital images at multiple focal planes of the entire slide, which demands long scanning time, complex and expensive scanning systems, and huge storage capacity. Aims: Here we report a unique imaging method that can acquire cytologic images efficiently and computationally render all-in-focus digital images that are highly compact. Methods and material: This method applies a metric-based digital refocusing to microscopy data collected with a Fourier ptychographic microscope (FPM). The digitally refocused patches of images are then synthesized into an all-in-focus image. Results: We report all-in-focus FPM results of thyroid fine needle aspiration (FNA) cytology samples, demonstrating our method\u27s ability to overcome the height variance of 30 μm caused by cell aggregation, and rendering images at high resolution (corresponds to a standard microscope with objective NA of 0.75) and that are all-in-focus. Conclusions: This technology is applicable to standard microscopes, and we believe can have an impact on diagnostic accuracy as well as ease and speed of diagnosing challenging specimens. While we focus on cytology slides here, we anticipate this technology\u27s advantages will translate well for histology applications. This technique also addresses the issue of remote rapid evaluation of cytology preparations. Finally, we believe that by resolving the focus heterogeneity issues in standard digital images, this technique is a critical advance for applying machine learning to cytology specimens

Persistent Helicobacter pylori Specific Th17 Responses in Patients with Past H. pylori Infection Are Associated with Elevated Gastric Mucosal IL-1β

10.1371/journal.pone.0039199PLoS ONE76

All-in-focus fine needle aspiration biopsy imaging based on Fourier ptychographic microscopy

Context: Cytology is the study of whole cells in diagnostic pathology. Unlike standard histologic thinly sliced specimens, cytologic preparations consist of preparations of whole cells where cells commonly cluster and aggregate. As such, cytology preparations are generally much thicker than histologic slides, resulting in large patches of defocus when examined under the microscope. A diagnostic aggregate of cells often cannot be viewed in focus together, requiring pathologists to continually manipulate the focal plane, complicating the task of accurately assessing the entire cellular aggregate and thus in making a diagnosis. Further, it is extremely difficult to acquire useful uniformly in-focus digital images of cytology preparations for applications such as remote diagnostic evaluations and artificial intelligence models. The predominant current method to address this issue is to acquire digital images at multiple focal planes of the entire slide, which demands long scanning time, complex and expensive scanning systems, and huge storage capacity. Aims: Here we report a unique imaging method that can acquire cytologic images efficiently and computationally render all-in-focus digital images that are highly compact. Methods and material: This method applies a metric-based digital refocusing to microscopy data collected with a Fourier ptychographic microscope (FPM). The digitally refocused patches of images are then synthesized into an all-in-focus image. Results: We report all-in-focus FPM results of thyroid fine needle aspiration (FNA) cytology samples, demonstrating our method’s ability to overcome the height variance of 30 μm caused by cell aggregation, and rendering images at high resolution (corresponds to a standard microscope with objective NA of 0.75) and that are all-in-focus. Conclusions: This technology is applicable to standard microscopes, and we believe can have an impact on diagnostic accuracy as well as ease and speed of diagnosing challenging specimens. While we focus on cytology slides here, we anticipate this technology’s advantages will translate well for histology applications. This technique also addresses the issue of remote rapid evaluation of cytology preparations. Finally, we believe that by resolving the focus heterogeneity issues in standard digital images, this technique is a critical advance for applying machine learning to cytology specimens

COVID-19 post-vaccination lymphadenopathy: Report of cytological findings from fine needle aspiration biopsy

10.1002/dc.24863DIAGNOSTIC CYTOPATHOLOGY4912E467-E47

SOX11 expression in a case of papillary thyroid carcinoma with fibromatosis/fasciitis-like stroma containing BRAF c.1799_1801delTGA and CTNNB1 c.133T>C mutations

10.1007/s00428-019-02619-4VIRCHOWS ARCHIV4754519-52

Idiopathic Granulomatous Mastitis and Erythema nodosum - A Unifying Pathophysiology?

10.1111/ajd.13463AUSTRALASIAN JOURNAL OF DERMATOLOGY621E149-E15

Lipidomic Analysis of Archival Pathology Specimens Identifies Altered Lipid Signatures in Ovarian Clear Cell Carcinoma

Cancer metabolism is associated with the enhanced lipogenesis required for rapid growth and proliferation. However, the magnitude of dysregulation of diverse lipid species still requires significant characterization, particularly in ovarian clear cell carcinoma (OCCC). Here, we have implemented a robust sample preparation workflow together with targeted LC-MS/MS to identify the lipidomic changes in formalin-fixed paraffin-embedded specimens from OCCC compared to tumor-free ovarian tissue. We quantitated 340 lipid species, representing 28 lipid classes. We observed differential regulation of diverse lipid species belonging to several glycerophospholipid classes and trihexosylceramide. A number of unsaturated lipid species were increased in OCCC, whereas saturated lipid species showed a decrease in OCCC compared to the controls. We also carried out total fatty acid analysis and observed an increase in the levels of several unsaturated fatty acids with a concomitant increase in the index of stearoyl-CoA desaturase (SCD) in OCCC. We confirmed the upregulation of SCD (the rate-limiting enzyme for the synthesis of monounsaturated fatty acids) by immunohistochemistry (IHC) assays. Hence, by carrying out a mass spectrometry analysis of archival tissue samples, we were able to provide insights into lipidomic alterations in OCCC

The IL-17A response of CD4⁺ PBMCs to HP-pulsed APCs.

<p>The mean ± SEM (standard error of mean) has been depicted for measurements of IL-17A concentration in the supernatant of co-culture experiments performed as described in the legend for <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0039199#pone-0039199-g003" target="_blank">figure 3B</a>. CD4⁺ T cells were co-cultured with autologous HP-pulsed APCs either in the absence or presence of MHC Class II blocking antibody. The fold-decrease in response was calculated by dividing the supernatant concentration of IL-17A from the culture without MHC Class II blockade, by the IL-17A concentration in the culture with MHC Class II blockade. The mean fold-decrease was obtained for the indicated number of biological replicates in each group, and mean ± SEM has been reported in the table.</p

Elevated frequencies of cells that express IL-17A persist in individuals with past HP infection.

<p>(A) The percentage of CD3⁺CD8⁻CCR6⁺IL-17A⁺ cells as a function of CD3⁺CD8⁻ PBMCs was assessed by flow cytometry. PBMCs were activated with PMA and ionomycin for 5 hours in the presence of GolgiStop. Cells were stained for cell surface CD3, CD8, and CCR6, fixed, permeabilised, and stained for intracellular IL-17A. Th17 cells were defined as CCR6⁺IL-17A⁺ events within the CD3⁺CD8⁻ compartment. Representative flow cytometry plots of individuals from group A, P, and N have been depicted. (B) Scatter plot of CD3⁺CD8⁻CCR6⁺IL-17A⁺ cells as a percentage of CD3⁺CD8⁻ cells among PBMCs that had been stimulated with PMA and ionomycin. Group A (n = 44), group P (n = 47), and group N (n = 48). The median and interquartile ranges have been represented on the scatter plot as horizontal bars. (C) The frequency of CD3⁺CD8⁻CCR6⁺IL-17A⁺ events within the CD3⁺CD8⁻ compartment for individuals from group P divided according to years since HP treatment. 1 year (n = 13), 2–3 years (n = 9), 4–9 years (n = 7), and ≥10 years (n = 3). (D) Number of CD4⁺IL-17A⁺ cells per high powered field (HPF) in gastric biopsy samples. Immunofluorescence microscopy was performed on gastric biopsies obtained from 8 patients in group A, 17 patients in group P, 12 patients in group N. For each patient sample, ten HPFs were evaluated and the average number of CD4⁺IL-17A⁺ cells per HPF was represented on the scatter plot. (E) Number of CD4⁺IL-17A⁺ cells per HPF in samples from group P stratified according to years since HP treatment. 1 year (n = 3), 2–3 years (n = 5), 4–9 years (n = 1), and ≥10 years (n = 3). (F − I) Cytokine concentrations in clarified homogenate obtained from mechanically disrupted gastric biopsy samples were measured using the MILLIPLEX® xMAP® bead-based cytokine quantification assay. (F) IL-17A concentration in gastric biopsy samples obtained from patients in group A (n = 7), group P (n = 15), and group N (n = 9). (G) IL-17A concentration in gastric biopsy samples from group P individuals depicted in (F) who were further sub-grouped based on the presence (PC+) or absence (PC<b>−</b>) of histological evidence of pre-cancerous lesions (chronic atrophic gastritis or intestinal metaplasia) in the gastric mucosa. PC+ (n = 12), PC<b>−</b> (n = 3). (H) IFNγ concentration in gastric biopsy samples obtained from patients in group A (n = 7), group P (n = 15), and group N (n = 9). (I) IL-8 concentration in gastric biopsy samples obtained from patients in group A (n = 7), group P (n = 15), and group N (n = 9). NS: not significant, *p<0.05, **p<0.001, ***p<0.0001.</p