Biology subjects

Kusmartseva, I.

Publications and source records attributed to Kusmartseva, I..

15 recordsLinked to original sources

bioRxiv2026

Virtual multiplex staining of the pancreatic islets across type 1 diabetes progression using a Schroedinger bridge

Classical hematoxylin and eosin (H&E) staining enables review of tissue morphology but lacks information regarding the molecular state of cells. Immunohistochemical (IHC) techniques label specific proteins in tissue, allowing differentiation of relevant structures that may go undetectable in H&E. However, the IHC process is complex, expensive, and time-consuming, especially for multiplex IHC (mIHC) limiting its use in large cohorts. Stain conversion of H&E to IHC using generative artificial intelligence models such as generative adversarial networks (GANs) represent one solution to this problem. However, GANs are unstable during out of distribution sampling and are prone to hallucinations or mode collapse, limiting their accuracy in challenging image conversion tasks. To address this, the field has recently turned to diffusion models. Here, we introduce Schrodinger-bridge for Multiplex ImmunoLabel Estimation (SMILE). Unlike conventional diffusion models that map from source to target through an intermediate Gaussian noise, Schrodinger-bridge diffusion models skip this step and have been shown to better preserve structures during image translation. To test the performance of SMILE, we generated a large cohort of high-fidelity H&E-mIHC image pairs from pancreatic organ donors, targeting insulin, glucagon, and CD3. Our dataset well-sampled across type-1 diabetes status, pancreas anatomical location, age, and sex. Using this cohort, we demonstrate the superiority of SMILE compared to GANs via a comprehensive evaluation framework incorporating texture, distribution, and antibody-specific metrics, as well as blinded pathologist reviews. We further confirmed the ability of SMILE to generate accurate mIHC images from H&Es generated at an external site, to perform whole slide image conversion, and to generate realistic three-dimensional maps of the pancreatic islets in non-diabetic, auto-antibody positive, and type-1 diabetic donor tissue. Finally, we performed stain conversion of paired H&E to HER2 and Ki67 images in breast cancer, confirming the superiority of SMILE in diverse stain conversion applications. Collectively, this framework provides a scalable pipeline for high-throughput proteomic inference from archival H&Es, providing transformative potential for pancreatic research and digital pathology.

bioinformatics↗

bioRxiv2025

3D imaging of human pancreas suggests islet size and endocrine composition influence their loss in type 1 diabetes

A high-definition description of pancreatic islets would prove beneficial for understanding the pathophysiology of type 1 diabetes (T1D), yet significant knowledge voids exist in terms of their size, endocrine cell composition, and number in both health and disease. Here, 3-dimensional (3D) analyses of pancreata from control persons without diabetes (ND) revealed heretofore underappreciated frequencies (approximately 50%) of insulin-positive (INS+) glucagon-negative (GCG-) islets. Non-diabetic individuals positive for a single Glutamic acid decarboxylase autoantibody (GADA+) yet at increased risk for disease consistently demonstrated endocrine features, including islet volume and cell composition, closely resembling the age-matched ND controls. In contrast, pancreata from individuals with short-duration T1D demonstrated significantly reduced islet density and a dramatic loss of INS+GCG- islets with preservation of large INS+GCG+ islets. The size and cellular composition of pancreatic islets may, therefore, represent influential factors that impact {beta}-cell loss during T1D disease progression. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=127 SRC="FIGDIR/small/654045v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@1d2fdeaorg.highwire.dtl.DTLVardef@11c43edorg.highwire.dtl.DTLVardef@af8a82org.highwire.dtl.DTLVardef@ae8c58_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LI3D imaging of non-diabetic (ND) pancreas suggests up to 50% of INS+ islets lack GCG C_LIO_LIINS+GCG- islets are typically small-sized in ND and preferentially lost in T1D C_LIO_LIIn T1D pancreas, INS+ beta cells are preserved in large INS+GCG+ islets C_LIO_LIIslets from GADA+ individuals appear similar to ND for size and {beta}-to- cell ratios C_LI

pathology↗

bioRxiv2025

Small things matter: Lack of extra-islet beta cells in Type 1 diabetes

Recent 3D analyses reported abundant, small beta-cell-rich endocrine objects (EOs) in the human pancreas. Here, we assessed morphological parameters of >262,000 EOs in pancreas sections from 220 donors with or without type 1 diabetes (T1D), ranging in age and disease duration. We observe many insulin (Ins)+/glucagon (Gluc)-EOs in donors without diabetes. Their relative contribution to the total endocrine area is greatest in early life (0-2y) but reduces thereafter. Strikingly, we show the virtual absence of Ins+Gluc- EOs in individuals with T1D, where only the medium and large EOs retain beta cells. We also report a lower EO density in T1D, especially in individuals diagnosed in early life. These findings suggest that extra-islet beta cells are impacted in the development of T1D, and their early loss is a characteristic feature. This new understanding has important implications for defining beta-cell mass, which may inform future screening and treatment strategies in T1D. HighlightsO_LIThe present extensive 2D studies confirm and extend recent 3D analyses of human pancreata, demonstrating that 50% of endocrine objects (EOs) are much smaller than classical islets of Langerhans and consist predominantly of beta cells (Ins+). C_LIO_LISmall insulin+ EOs comprise the largest proportion of the total endocrine area in early childhood and persist throughout the life course in donors without diabetes. C_LIO_LIThere is a shift towards larger EO size with increasing age, with the most pronounced changes in size occurring in the first few years of life. C_LIO_LISmall Ins+ EOs are virtually absent in individuals with type 1 diabetes, while the persisting EOs with beta cells are larger, suggesting a selective destruction of beta cells in small EOs. C_LIO_LIDevelopment of type 1 diabetes, particularly at an early age, is associated with fewer larger EOs in adulthood. This implies that the lack or early destruction of small Ins+ EOs may be detrimental to the generation of larger EOs. C_LI

pathology↗

bioRxiv2025

Integration of nuclear morphology and 3D imaging to profile cellular neighborhoods

Nuclear morphology is an indicator of cellular function and disease states, as changes in nuclear size, shape, and texture often reflect underlying disease-related genetic, epigenetic, and microenvironmental alterations. For disease diagnosis, nuclear segmentation performed in 2D hematoxylin and eosin (H&E)-stained tissue sections has long represented the gold standard. However, recent advances in three-dimensional (3D) histology, which provide a more biologically accurate representation of the spatial heterogeneity of human microanatomy, has led to improved understandings of disease pathology. Yet challenges remain in the development of scalable and computationally efficient pipelines for extracting and interpreting nuclear features in 3D space. 2D histology neglects crucial spatial information, such as 3D connectivity, morphology, and rare events missed by sparser sampling. Here, through extension of the CODA platform, we integrate 3D imaging with nuclear segmentation to analyze nuclear morphological features in human tissue. Analysis of 3D tissue microenvironments uncovered critical changes in 3D morphometric heterogeneity. Additionally, it enables the spatial characterization of immune cell distribution in relation to tissue structures, such as variations in leukocyte density near pancreatic ducts and blood vessels of different sizes. This approach provides a more comprehensive understanding of tissue and nuclear structures, revealing spatial patterns and interactions that are critical for disease progression.

bioinformatics↗

bioRxiv2025

Integrated histopathology of the human pancreas throughout stages of type 1 diabetes progression

Type 1 diabetes (T1D) is a progressive autoimmune condition that culminates in the loss of insulin-producing beta cells. Pancreatic histopathology provides essential insights into disease initiation and progression yet an integrated perspective of in situ pathogenic processes is lacking due to limited sample availability, the dispersed nature of anatomical lesions, and often restricted analytical dimensionality. Here, we combined multiplexed immunostaining, high-magnification whole-slide imaging, digital pathology, and semi-automated image analysis strategies to interrogate pancreatic tail and head regions obtained from organ donors across T1D stages including at-risk and at-onset cases. Deconvolution of architectural features, endocrine cell composition, immune cell burden, and spatial relations of [~]25,000 islets revealed a series of novel histopathological correlates especially in the prodromal disease stage preceding clinical T1D. Altogether, our comprehensive "single-islet" analyses permit the reconstruction of a revised natural T1D history with implications for further histopathological investigations, considerations of pathogenetic modalities, and therapeutic interventions.

immunology↗

bioRxiv2025

Imaging mass cytometry reveals early β-cell dysfunction and changes in immune signatures during type 1 diabetes progression in human pancreata

The natural history and pathogenesis of type 1 diabetes, particularly during the autoantibody- positive stages preceding clinical onset, are not well understood, in part, due to limited availability of human pancreatic samples. Here, we studied 88 organ donors, including 28 single autoantibody-positive and 10 multiple autoantibody-positive donors, by imaging mass cytometry. Approximately 10,000 islets and 16 million single-cells were spatially analyzed using 79 antibodies revealing both {beta}-cell states and the islet-immune interface. We identified IAPP loss from {beta}-cells as an indicator of pre-clinical disease. Alterations in Interferon signatures and downregulation across lineage and functional markers, including markers of endoplasmic reticulum stress, were characteristic of recent-onset disease. Further, in single autoantibody- positive donors, we identified pro-inflammatory myeloid cells and PD1+ memory CD4+ T cells, and in multiple autoantibody-positive samples, found islet-specific and exhausted-like ebector CD8+ T cells. Multiple immune cell subtypes were associated with young age, disease severity and insulitis. This dataset is a major step toward creation of a multi-modal type 1 diabetes disease atlas that will be useful for identifying potential drug targets and association of disease features with clinical co-variates and trial outcomes.

immunology↗

bioRxiv2025

Single-cell multiome and spatial profiling reveals pancreas cell type-specific gene regulatory programs driving type 1 diabetes progression

Cell type-specific regulatory programs that drive type 1 diabetes (T1D) in the pancreas are poorly understood. Here we performed single nucleus multiomics and spatial transcriptomics in up to 32 non-diabetic (ND), autoantibody-positive (AAB+), and T1D pancreas donors. Genomic profiles from 853,005 cells mapped to 12 pancreatic cell types, including multiple exocrine sub- types. Beta, acinar, and other cell types, and related cellular niches, had altered abundance and gene activity in T1D progression, including distinct pathways altered in AAB+ compared to T1D. We identified epigenomic drivers of gene activity in T1D and AAB+ which, combined with genetic association, revealed causal pathways of T1D risk including antigen presentation in beta cells. Finally, single cell and spatial profiles together revealed widespread changes in cell-cell signaling in T1D including signals affecting beta cell regulation. Overall, these results revealed drivers of T1D progression in the pancreas, which form the basis for therapeutic targets for disease prevention.

genomics↗

bioRxiv2024

Beta cell dysfunction occurs independently of insulitis in type 1 diabetes pathogenesis

The loss of insulin secretory function associated with type 1 diabetes (T1D) is attributed to the immune-mediated destruction of beta cells. Yet, at onset of T1D, patients often have a significant beta cell mass remaining while T cell infiltration of pancreatic islets is sporadic. Thus, we investigated the hypothesis that the remaining beta cells in T1D are largely dysfunctional using live human pancreas tissue slices prepared from organ donors with recently diagnosed T1D. Beta cells in slices from donors with T1D had significantly diminished Ca2+ mobilization and insulin secretion responses to glucose. Beta cell function was equally impaired in T cell-infiltrated and non-infiltrated islets. Fixed tissue staining and gene expression profiling of laser-capture microdissected islets revealed significant decreases of proteins and genes in the glucose stimulus secretion coupling pathway. From these data, we posit that functional defects occur in the remaining mass of beta cells during human T1D pathogenesis.

physiology↗

bioRxiv2024

Mapping histological and functional maturation of human endocrine pancreas across early postnatal periods

Human endocrine cell differentiation and islet morphogenesis play critical roles in determining islet cell mass and function, but the events and timeline of these processes are incompletely defined. To better understand early human islet cell development and maturation, we collected 115 pediatric pancreata and mapped morphological and spatiotemporal changes from birth through the first ten years of life. Using quantitative analyses and a combination of complementary tissue imaging approaches, including confocal microscopy and whole-slide imaging, we developed an integrated model for endocrine cell formation and islet architecture, including endocrine cell type heterogeneity and abundance, endocrine cell proliferation, and islet vascularization and innervation. We also assessed insulin and glucagon secretory profiles in isolated islet preparations from pediatric donors aged 2 months to 10 years and found a temporal difference in the maturation of insulin secretion compared to glucagon secretion. This comprehensive summary of postnatal and pediatric pancreatic islet development provides a framework for future studies and integration of emerging genetic and genomic data related to islet biology and diabetes risk.

developmental biology↗

bioRxiv2024

ENTPD3-specific CAR Regulatory T cells for Local Immune Control in T1D

Despite advances in Type 1 Diabetes (T1D) management such as hybrid closed loop systems, patients still face significant morbidity, reduced life expectancy, and impaired glucose regulation compared to healthy individuals or those with pancreas transplants. Here we developed beta cell-specific Chimeric Antigen Receptors (CAR) targeting the antigen ectonucleoside triphosphate diphosphohydrolase 3 (ENTPD3) using a novel cell-based phage display methodology. ENTPD3 is highly expressed on beta cells of both early and progressed T1D patients. ENTPD3 CAR regulatory T cells (Tregs) homed, expanded and persisted in pancreatic islets in a T1D mouse model (NOD) and completely prevented disease progression. Human ENTPD3 CAR Tregs displayed a stable regulatory phenotype, strong activation, and suppression. Importantly, ENTPD3 CAR T cells recognised and were fully activated by human islets. This approach holds great promise as a durable treatment option for patients with prediabetes, new-onset diabetes, or those undergoing beta cell replacement therapy.

immunology↗

bioRxiv2021

3D-mapping of human lymph node and spleen reveals integrated neuronal, vascular, and ductal cell networks

The spleen and lymph node represent important hubs for both innate and adaptive immunity1,2. Herein, we map immune, endothelial, and neuronal cell networks within these tissues from "normal"/non-diseased organ donors, collected through the NIH Human BioMolecular Atlas Program (HuBMAP)3, using highly multiplexed CODEX (CO-Detection by indEXing) imaging and 3D light sheet microscopy of cleared tissues. Building on prior reports4-6, we observed the lymph node subcapsular sinus expressing podoplanin, smooth muscle actin, and LYVE1. In the spleen, LYVE1 was expressed by littoral cells lining venous sinusoids, whereas podoplanin was restricted to arteries and trabeculae. 3D visualization of perivascular innervation revealed a subset of axonal processes expressing choline acetyl transferase in both tissues, in contrast with prior literature on human spleen7. We further report our novel observations regarding the distinct localization of GAP43 and {beta}3-tubulin within the vascular anatomy of both lymph node and spleen, with Coronin-1A+ cells forming a dense cluster around {beta}3-tubulin positive GAP43 low/negative segments of large vessels in spleen. These data provide an unprecedented 2D and 3D visualization of cellular networks within secondary lymphoid tissues, laying the groundwork for future disease-specific and system-wide studies of neural regulation of immunity in human lymphatics.

immunology↗

bioRxiv2021

Single-cell analysis of the human pancreas in type 2 diabetes using multi-spectral imaging mass cytometry

Type 2 diabetes mellitus (T2D) is a chronic age-related disorder characterized by hyperglycemia due to the failure of pancreatic beta cells to compensate for increased insulin demand, typically associated with peripheral insulin resistance. However, despite decades of research, the pathogenic mechanisms underlying T2D remain poorly defined. Imaging mass cytometry (IMC) enables multiplexed assessment of the abundance and subcellular localization of multiple proteins on the same tissue section. Herein, we utilized IMC with a panel of 34 antibodies to quantify key markers of pancreatic exocrine, islet, and immune cells as well as stromal components. We employed this panel to analyze over 2.1 million cells from 16 pancreata obtained from donors with T2D and 13 pancreata from age similar non-diabetic controls. In the T2D pancreata, we observed significant alterations in islet architecture, endocrine cell composition, and surprisingly immune cell constituents. Thus, both HLA-DR positive CD8 T cells and macrophages were enriched intra-islet in the T2D pancreas. These efforts demonstrate the utility of IMC to investigate complex events at the cellular level in order to provide new insights into the pathophysiology of T2D.

molecular biology↗

bioRxiv2020

ACE2 and SARS-CoV-2 Expression in the Normal and COVID-19 Pancreas

Diabetes is associated with increased mortality from Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). Given literature suggesting a potential association between SARS-CoV-2 infection and diabetes induction, we examined pancreatic expression of the key molecule for SARS-CoV-2 infection of cells, angiotensin-converting enzyme-2 (ACE2). Specifically, we analyzed five public scRNAseq pancreas datasets and performed fluorescence in situ hybridization, Western blotting, and immunolocalization for ACE2 with extensive reagent validation on normal human pancreatic tissues across the lifespan, as well as those from coronavirus disease 2019 (COVID-19) patients. These in silico and ex vivo analyses demonstrated pancreatic expression of ACE2 is prominent in pancreatic ductal epithelium and the microvasculature, with rare endocrine cell expression of this molecule. Pancreata from COVID-19 patients demonstrated multiple thrombotic lesions with SARS-CoV-2 nucleocapsid protein expression primarily limited to ducts. SARS-CoV-2 infection of pancreatic endocrine cells, via ACE2, appears an unlikely central pathogenic feature of COVID-19 as it relates to diabetes.

pathology↗

bioRxiv2020

Pancreatlas: a comprehensive and interactive image atlas of the human pancreas in health and disease

Human tissue phenotyping generates complex spatial information from numerous imaging modalities, yet images typically become static figures for publication and original data and metadata are rarely available. While comprehensive image maps exist for some organs, most resources have limited support for multiplexed imaging or have non-intuitive user interfaces. Therefore, we built a Pancreatlas resource that integrates several technologies into a novel interface, allowing users to access richly annotated web pages, drill down to individual images, and deeply explore data online. The current version of Pancreatlas contains over 800 unique images acquired by whole-slide scanning, confocal microscopy, and imaging mass cytometry, and is available at https://www.pancreatlas.org. To create this human pancreas-specific biological imaging resource, we developed a React-based web application and Python-based application programming interface, collectively called Flexible Framework for Integrating and Navigating Data (FFIND), which can be adapted beyond Pancreatlas to meet countless imaging or other structured data management needs.

cell biology↗

bioRxiv2018

Detection of enterovirus protein and RNA in multiple tissues from nPOD organ donors with type 1 diabetes

Epidemiological studies have shown an association between enterovirus (EV) infections and type 1 diabetes (T1D), and EV protein has been detected in the pancreatic islets of T1D patients. Here we correlated the detection of EVs in lymphoid tissues (spleen and pancreatic lymph nodes) and small intestinal mucosa to the virus detection in the pancreas of T1D, autoantibody-positive (aab+) and non-diabetic control organ donors of the Network for Pancreatic Organ Donors with Diabetes (nPOD) study. Formalin-fixed paraffin-embedded tissue samples were screened for insulin and EV protein using immunohistochemistry, and frozen tissue for EV genome using RT-PCR. The presence of EV protein in the pancreatic islets correlated with the presence of insulin-positive cells. Altogether 62 % of T1D and aab+ donors were positive for EV protein in pancreatic islets (only insulin-positive donors included), 40 % in duodenum and 32 % in spleen, compared to 33 %, 14 %, and 27 % of non-diabetic controls. Pancreatic lymph nodes were positive for EV protein in 60 % of T1D and aab+ cases. T1D and aab+ donors were more frequently VP1-positive in multiple organs than control donors (39 % vs. 11 %; including only insulin-positive donors). EV RNA was found in selected donors and from multiple tissue types except for duodenum, and individual T1D and aab+ donors were EV RNA-positive in multiple organs. The role of extra-pancreatic organs and their interplay with EV in T1D pathogenesis remains to be solved, but we hypothesize that these organs may serve as a reservoir for the virus which may reside in these tissues in a slow-replicating persistent form.

epidemiology↗