Biology subjects

Kuehnel, M.

Publications and source records attributed to Kuehnel, M..

5 recordsLinked to original sources

bioRxiv2024

Aberrant and Ectopic Cell Populations of the Fibrotic Pushing Border in Restrictive Allograft Syndrome after Lung Transplantation

RationaleRestrictive allograft syndrome (RAS) is a major cause of mortality in patients following lung transplantation due to rapid progressive fibrosis in the pulmonary graft. We have only limited knowledge of the cellular and molecular mechanisms that characterize the fibrosis in the RAS lung. ObjectiveTo elucidate cellularly-resolved transcriptomic and histologic characteristics of the structural cells in human RAS lungs. MethodsSingle-nuclei RNA-sequencing was performed in peripheral lung tissues from 15 RAS patients undergoing lung re-transplantation, and from 9 healthy control lungs. Findings were validated and complemented by various histologic techniques, including immunofluorescence, RNAscope, combined Elastica van Gieson-immunohistochemistry stains, and micro-CT scans. Measurement and Main resultsDifferential gene expression analysis of our single-nuclei RNA-sequencing data revealed in human RAS lungs previously undescribed and uniquely distributed aberrant basaloid cells, ectopic COL15A1+ vascular endothelial cells, and CTHRC1+ fibrotic fibroblasts, all first characterized in idiopathic pulmonary fibrosis (IPF). In contrast to IPF, RAS lacks the cellular equivalent of bronchiolization. Histologic stains confirmed our transcriptomic discoveries and disclosed distinctive distribution patterns: Aberrant basaloid cells are primarily localized at the edge of the fibrotic pushing border, forming together with the juxtaposed CTHRC1+ fibrotic fibroblasts the fibrotic niche of alveolar fibroelastosis (AFE), the histopathological hallmark in RAS lungs. On the endothelial side, PRX+ alveolar microvasculature is lost in AFE areas. Micro-CT scans revealed that blood supply, now facilitated by expanded and ectopic COL15A1+ VE cells, changes from pulmonary to systemic perfusion. Last, our data reveals potential therapeutically-modifiable expression patterns in RAS, including genes coding for the integrin subunits v{beta}6, activators of TGF{beta}. ConclusionConsidering the marked clinical, histologic and etiologic dissimilarities of RAS and IPF, our snRNAseq study revealed a surprising general principle of cellular and molecular pathogenesis in the fibrosing lung: the entity-spanning composition of the fibrotic niche by a) aberrant basaloid cells localized at the fibrotic pushing border, b) ectopic COL15A1+ vascular ECs and c) effector CTHRC1+ fibrotic fibroblasts. This general principle justifies a flexible but cellular pathogenesis-guided transferability of potential therapeutic approaches between progressive fibrotic lung diseases.

cell biology↗

bioRxiv2024

Cell therapy with human iPSC-derived cardiomyocyte aggregates leads to efficient engraftment and functional recovery after myocardial infarction in non-human primates

BackgroundFunctionally coupled large myocardial grafts and a remarkable improvement of heart function in nonhuman primate models of myocardial infarction have been reported after transplantation of human embryonic stem cell-derived cardiomyocytes at relatively high numbers of up to 109 single cell cardiomyocytes - a dose equivalent to total cell loss after myocardial infarction in [~]10 times larger human hearts. To overcome apparent limitations associated with the application of single cells, this pre-clinical study investigated the injection of cardiomyocyte aggregates instead. MethodsHuman iPSC-derived cardiomyocyte aggregates were produced in scalable suspension culture. Intramyocardial injection of the aggregates into cynomolgus monkey hearts was conducted two weeks after myocardial infarction induced by permanent coronary artery ligation. Human cell engraftment was assessed after two weeks or three months; functional analyses included continuous telemetric ECG recording and repeated cardiac MRI assessment in comparison to sham treated animals. ResultsTreatment with cell numbers as low as 5 x 107 resulted in efficient structural engraftment. Notably, the degree of heart function recovery in vivo seemed to correlate with the contractility of the applied cardiomyocytes tested by parallel experiments in vitro. Graft-induced non-life-threatening arrhythmias were transient and decreased considerably during the three months follow-up. ConclusionsTransplantation of human iPSC-derived cardiomyocyte aggregates yielded comparable results to the reported application of higher numbers of single cell cardiomyocytes from human ESC, suggesting that the application of cardiomyocyte aggregates facilitates cell therapy development by reducing cell production costs and clinical risks associated with the administration of relatively high cell numbers. Clinical PerspectiveWhat is new? O_LIIn contrast to previously applied single cells, human iPSC-derived cardiomyocyte aggregates (hiCMAs) were transplanted in a non-human primate (NHP) model of MI, to reduce the required cell dose, promote myocardial retention of the graft, and limit the risks for adverse effects. Such low-dose treatment with almost pure ventricular cardiomyocytes produced under GMP-compliant conditions, resulted in the formation of relative large, structurally integrated human grafts in NHP hearts. C_LIO_LITransient non-life-threatening arrhythmias associated with intramyocardial cell transplantation decreased considerably during the three months follow-up. C_LIO_LIA remarkable recovery of left ventricular function was observed. This recovery notably correlated with the in vitro contractility of transplanted cardiomyocyte batches tested in bioartificial cardiac tissues (BCTs), underlining the relevance of a suitable potency assay. C_LI What are the clinical implications? O_LIIntra-myocardial injection of hiCMAs is a promising treatment modality for the recovery of contractile function after MI; their advanced production, storage and testing revealed in the study facilitate the clinical translation of hiPSC-based heart repair. C_LIO_LIThe need for relatively low numbers of cardiomyocytes produced through advanced protocols for scalable suspension culture reduces production costs of adequate cell batches, thereby increasing treatment availability. In vitro testing of the produced cell batches is required to ensure treatment efficacy. C_LIO_LIClinical hiCMA injection can be considered reasonably safe, however, pharmacological prevention and treatment of arrhythmias is required and temporary implantation of a cardioverter-defibrillator (ICD) could be considered. C_LI

cell biology↗

bioRxiv2023

Spatial Transcriptomic Characterization of Novel Pathologic Niches in IPF

An unmet medical need persists in Idiopathic Pulmonary fibrosis (IPF), for which treatments additional to anti-fibrotic therapy are needed. Single cell RNA sequencing (scRNA-seq) has advanced our understanding of IPF with cell type-specific insights but lacks cellular tissue context. Spatial transcriptomics addresses this by providing spatially resolved gene expression, enabling gene and cell type localization within the tissue environment. We profiled IPF and control patient lung tissue sections using spatial transcriptomics and combined the data with an atlas of integrated IPF scRNA-seq datasets. Through computational analysis, we identified three disease-associated pathologic niches with unique cellular composition / localization and analyzed their cell-cell communication. We identified the Fibrotic niche, comprising Myofibroblasts and Aberrant Basaloid cells, preferentially located around airways and close to the Airway Macrophage niche in the lumen, containing SPP1+ Macrophages. We also identified the Immune niche, distinct foci of lymphoid cells in fibrotic tissue, surrounded by remodeled endothelial vessels. TEASERSpatial transcriptomics localizes genes and cell types in the tissue and identifies pathological cellular niches in IPF and control lungs.

systems biology↗

bioRxiv2023

TIGIT drives the immunosuppressive environment by downregulation of metalloproteinases MMP2 and MMP14 in perihilar cholangiocarcinoma.

BackgroundCheckpoint blockade in cholangiocarcinoma (CCA) is promising; however, little is known about the response to treatment in CCA subtypes. In this study, we investigated the spatial immune environment in combination with checkpoint expression in perihilar CCA (pCCA). Materials & MethodsThe levels of checkpoint molecules (PD-1, PD-L1, PD-L2, LAG-3, ICOS, TIGIT, TIM-3, and CTLA-4), macrophages (CD68), and T cells (CD4 and CD8) were assessed by multiplex immunofluorescence (mIF) in 50 patients. We investigated the transcriptomic profile using the NanoString Cancer Progression Panel, and validation was performed by mIF on tissue sections from 24 patients. ResultsThe expression of checkpoint molecules TIGIT, CTLA-4, and LAG-3 alone and in combination with other checkpoint molecules was more abundant in the Central Tumor (CT) and Invasive Margin (IM) than in peritumoral tissue (PT) (CD4 and CD8 TIGIT p<0.0001 for both CD4 and CD8 CTLA-4, p<0.0001 and p < 0.001, respectively, and CD8 LAG-3 p < 0.05). MMP2 and MMP14 were differentially expressed in patients with high TIGIT expression. ConclusionThe immune environment in pCCA is characterized by the expression of multiple checkpoints, demonstrating the complexity of ICI treatment. High TIGIT expression drives an immunosuppressive environment by modulating the extracellular matrix. Future clinical trials in pCCA could consider TIGIT as a therapeutically relevant target for (combination) treatment.

cancer biology↗

bioRxiv2022

Hamsters are a model for COVID-19 alveolar regeneration mechanisms: an opportunity to understand post-acute sequelae of SARS-CoV-2

A relevant number of coronavirus disease 2019 (COVID-19) survivors suffers from post-acute sequelae of severe acute respiratory syndrome coronavirus 2 (PASC). Current evidence suggests a dysregulated alveolar regeneration in COVID-19 as a possible explanation for respiratory PASC symptoms, a phenomenon which deserves further investigation in a suitable animal model. This study investigates morphological, phenotypical and transcriptomic features of alveolar regeneration in SARS-CoV-2 infected Syrian golden hamsters. We demonstrate that CK8+ alveolar differentiation intermediate (ADI) cells occur following SARS-CoV-2-induced diffuse alveolar damage. A subset of ADI cells shows nuclear accumulation of TP53 at 6- and 14-days post infection (dpi), indicating a prolonged arrest in the ADI state. Transcriptome data show the expression of gene signatures driving ADI cell senescence, epithelial-mesenchymal transition, and angiogenesis. Moreover, we show that multipotent CK14+ airway basal cell progenitors migrate out of terminal bronchioles, aiding alveolar regeneration. At 14 dpi, presence of ADI cells, peribronchiolar proliferates, M2-type macrophages, and sub-pleural fibrosis is observed, indicating incomplete alveolar restoration. The results demonstrate that the hamster model reliably phenocopies indicators of a dysregulated alveolar regeneration of COVID-19 patients. The results provide important information on a translational COVID-19 model, which is crucial for its application in future research addressing pathomechanisms of PASC and in testing of prophylactic and therapeutic approaches for this syndrome.

pathology↗