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Image-based cytometry faces challenges due to technical variations arising from different experimental batches and conditions, such as differences in instrument configurations or image acquisition protocols, impeding genuine biological interpretation of cell morphology. Existing solutions, often necessitating extensive pre-existing data knowledge or control samples across batches, have proved limited, especially with complex cell image data. To overcome this, "Cyto-Morphology Adversarial Distillation" (CytoMAD), a self-supervised multi-task learning strategy that distills biologically relevant cellular morphological information from batch variations, is introduced to enable integrated analysis across multiple data batches without complex data assumptions or extensive manual annotation. Unique to CytoMAD is its "morphology distillation", symbiotically paired with deep-learning image-contrast translation-offering additional interpretable insights into label-free cell morphology. The versatile efficacy of CytoMAD is demonstrated in augmenting the power of biophysical imaging cytometry. It allows integrated label-free classification of human lung cancer cell types and accurately recapitulates their progressive drug responses, even when trained without the drug concentration information. CytoMAD  also allows joint analysis of tumor biophysical cellular heterogeneity, linked to epithelial-mesenchymal plasticity, that standard fluorescence markers overlook. CytoMAD can substantiate the wide adoption of biophysical cytometry for cost-effective diagnosis and screening.

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OBJECTIVE: Lung cancers that present as radiographic subsolid nodules represent a subtype with distinct biological behavior and outcomes. The objective of this document is to review the existing literature and report consensus among a group of multidisciplinary experts, providing specific recommendations for the clinical management of subsolid nodules. METHODS: The American Association for Thoracic Surgery Clinical Practice Standards Committee assembled an international, multidisciplinary expert panel composed of radiologists, pulmonologists, and thoracic surgeons with established expertise in the management of subsolid nodules. A focused literature review was performed with the assistance of a medical librarian. Expert consensus statements were developed with class of recommendation and level of evidence for each of 4 main topics: (1) definitions of subsolid nodules (radiology and pathology), (2) surveillance and diagnosis, (3) surgical interventions, and (4) management of multiple subsolid nodules. Using a modified Delphi method, the statements were evaluated and refined by the entire panel. RESULTS: Consensus was reached on 17 recommendations. These consensus statements reflect updated insights on subsolid nodule management based on the latest literature and current clinical experience, focusing on the correlation between radiologic findings and pathological classifications, individualized subsolid nodule surveillance and surgical strategies, and multimodality therapies for multiple subsolid lung nodules. CONCLUSIONS: Despite the complex nature of the decision-making process in the management of subsolid nodules, consensus on several key recommendations was achieved by this American Association for Thoracic Surgery expert panel. These recommendations, based on evidence and a modified Delphi method, provide guidance for thoracic surgeons and other medical professionals who care for patients with subsolid nodules.

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INTRODUCTION: Lung transplantation is an established treatment for patients with end-stage lung disease. However, ischaemia reperfusion injury remains a barrier to achieving better survival outcomes. Here, we aim to investigate the metabolomic and transcriptomic profiles in human lungs before and after reperfusion, to identify mechanisms relevant to clinical outcome. METHODS: We analysed 67 paired human lung tissue samples collected from 2008 to 2011, at the end of cold preservation and 2 hours after reperfusion. Gene expression analysis was performed with R. Pathway analysis was conducted with Ingenuity Pathway Analysis. MetaboAnalyst and OmicsNet were used for metabolomics analysis and omics data integration, respectively. Association of identified metabolites with transplant outcome was investigated with Kaplan-Meier estimate and Cox proportional hazard models. RESULTS: Activation of energy metabolism and reduced antioxidative biochemicals were found by metabolomics. Upregulation of genes related to cytokines and inflammatory mediators, together with major signalling pathways were revealed by transcriptomics. Purine metabolism was identified as the most significantly enriched pathway at reperfusion, based on integrative analysis of the two omics data sets. Elevated expression of purine nucleoside phosphorylase (PNP) could be attributed to activation of multiple transcriptional pathways. PNP catabolised reactions were evidenced by changes in related metabolites, especially decreased levels of inosine and increased levels of uric acid. Multivariable analyses showed significant association of inosine and uric acid levels with intensive care unit length of stay and ventilation time. CONCLUSION: Oxidative stress, especially through purine metabolism pathway, is a major metabolic event during reperfusion and may contribute to the ischaemia reperfusion injury of lung grafts.

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BACKGROUND: The Omicron BA.2 sublineage has replaced BA.1 worldwide and has comparable levels of immune evasion to BA.1. These observations suggest that the increased transmissibility of BA.2 cannot be explained by the antibody evasion. METHODS: Here, we characterized the replication competence and respiratory tissue tropism of three Omicron variants (BA.1, BA.1.1, BA.2), and compared these with the wild-type virus and Delta variant, in human nasal, bronchial and lung tissues cultured ex vivo. FINDINGS: BA.2 replicated more efficiently in nasal and bronchial tissues at 33°C than wild-type, Delta and BA.1. Both BA.2 and BA.1 had higher replication competence than wild-type and Delta viruses in bronchial tissues at 37°C. BA.1, BA.1.1 and BA.2 replicated at a lower level in lung parenchymal tissues compared to wild-type and Delta viruses. INTERPRETATION: Higher replication competence of Omicron BA.2 in the human upper airway at 33°C than BA.1 may be one of the reasons to explain the current advantage of BA.2 over BA.1. A lower replication level of the tested Omicron variants in human lung tissues is in line with the clinical manifestations of decreased disease severity of patients infected with the Omicron strains compared with other ancestral strains. FUNDING: This work was supported by US National Institute of Allergy and Infectious Diseases and the Theme-Based Research Scheme under University Grants Committee of Hong Kong Special Administrative Region, China.

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The emergence of SARS-CoV-2 variants of concern with progressively increased transmissibility between humans is a threat to global public health. The Omicron variant of SARS-CoV-2 also evades immunity from natural infection or vaccines1, but it is unclear whether its exceptional transmissibility is due to immune evasion or intrinsic virological properties. Here we compared the replication competence and cellular tropism of the wild-type virus and the D614G, Alpha (B.1.1.7), Beta (B.1.351), Delta (B.1.617.2) and Omicron (B.1.1.529) variants in ex vivo explant cultures of human bronchi and lungs. We also evaluated the dependence on TMPRSS2 and cathepsins for infection. We show that Omicron replicates faster than all other SARS-CoV-2 variants studied in the bronchi but less efficiently in the lung parenchyma. All variants of concern have similar cellular tropism compared to the wild type. Omicron is more dependent on cathepsins than the other variants of concern tested, suggesting that the Omicron variant enters cells through a different route compared with the other variants. The lower replication competence of Omicron in the human lungs may explain the reduced severity of Omicron that is now being reported in epidemiological studies, although determinants of severity are multifactorial. These findings provide important biological correlates to previous epidemiological observations.

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Rapid and high-resolution histological imaging with minimal tissue preparation has long been a challenging and yet captivating medical pursuit. Here, the authors propose a promising and transformative histological imaging method, termed computational high-throughput autofluorescence microscopy by pattern illumination (CHAMP). With the assistance of computational microscopy, CHAMP enables high-throughput and label-free imaging of thick and unprocessed tissues with large surface irregularity at an acquisition speed of 10 mm2 /10 s with 1.1-µm lateral resolution. Moreover, the CHAMP image can be transformed into a virtually stained histological image (Deep-CHAMP) through unsupervised learning within 15 s, where significant cellular features are quantitatively extracted with high accuracy. The versatility of CHAMP is experimentally demonstrated using mouse brain/kidney and human lung tissues prepared with various clinical protocols, which enables a rapid and accurate intraoperative/postoperative pathological examination without tissue processing or staining, demonstrating its great potential as an assistive imaging platform for surgeons and pathologists to provide optimal adjuvant treatment.

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BACKGROUND: Necessity of flexible bronchoscopy (FB) examination as a routine preoperative work-up for peripheral clinical T1N0 subsolid lung cancer was unknown. METHODS: This was a prospective, multi-center clinical trial (NCT03591445). Patients with peripheral GGO nodules (GGNs) who were candidates for surgical resection were enrolled. FB examination was performed preoperatively. Surgical plan could be changed if any aberrant histologic and anatomic findings were detected by FB examination. Primary endpoint was the rate that surgical plan was changed by positive FB findings. Secondary endpoints were rate of positive FB findings and rate of procedural complications. RESULTS: Six hundred and fifteen patients with peripheral subsolid nodules detected by thoracic CT were enrolled. There were 187 (30.4%) male and 428 (69.6%) female patients, mean age was 54.85±10.41 y (range, 26-78). 262 (42.6%) patients had pure GGNs and 353 (57.4%) patients had part-solid nodules. Mean size of nodules was 13.87±6.37 mm (range, 5-30). FB examinations confirmed one (0.16%) adenocarcinoma, seven (1.14%) bronchial variations, one (0.16%) segmental bronchostenosis, one (0.16%) segmental bronchial occlusion and one (0.16%) bronchial inflammation. No complications of FB examinations occurred. 568 (92.35%) thoracoscopic and 47 (7.65%) open surgeries were performed. No established surgical plan was changed by positive FB findings. Final pathologies revealed 26 (4.2%) adenocarcinoma in situ (AIS), 240 (39%) minimal invasive adenocarcinomas (MIAs), 343 (55.8%) invasive adenocarcinomas (IADs), one (0.2%) adenosquamous cell carcinoma, one (0.2%) squamous cell carcinoma, two (0.3%) atypical adenoid hyperplasia and two (0.3%) inflammations. CONCLUSIONS: FB examination was unnecessary in the preoperative assessment of peripheral clinical T1N0 subsolid lung cancer.

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BACKGROUND: Normothermic ex vivo lung perfusion (EVLP) allows for functional assessment of donor lungs; thus has increased the use of marginal lungs for transplantation. To extend EVLP for advanced organ reconditioning and regenerative interventions, cellular metabolic changes need to be understood. We sought to comprehensively characterize the dynamic metabolic changes of the lungs during EVLP, and to identify strategies to improve EVLP. METHODS: Human donor lungs (n = 50) were assessed under a 4-hour Toronto EVLP protocol. EVLP perfusate was sampled at first (EVLP-1h) and fourth hour (EVLP-4h) of perfusion and were submitted for mass spectrometry-based untargeted metabolic profiling. Differentially expressed metabolites between the 2 timepoints were identified and analyzed from the samples of lungs transplanted post-EVLP (n = 42) to determine the underlying molecular mechanisms. RESULTS: Of the total 312 detected metabolites, 84 were up-regulated and 103 were down-regulated at EVLP-4h relative to 1h (FDR adjusted p < .05, fold change ≥ |1.1|). At EVLP-4h, markedly decreased energy substrates were observed, accompanied by the increase in fatty acid ß-oxidation. Concurrently, accumulation of amino acids and nucleic acids was evident, indicative of increased protein and nucleotide catabolism. The uniform decrease in free lysophospholipids and polyunsaturated fatty acids at EVLP-4h suggests cell membrane remodeling. CONCLUSIONS: Untargeted metabolomics revealed signs of energy substrate consumption and metabolic by-product accumulation under current EVLP protocols. Strategies to supplement nutrients and to maintain homeostasis will be vital in improving the current clinical practice and prolonging organ perfusion for therapeutic application to further enhance donor lung utilization.

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