El Shami Mohamad, Savani Milan R, Gattie Lauren C, Smith Bailey, Hicks William H, Rich Jeremy N, Richardson Timothy E, McBrayer Samuel K, Abdullah Kalil G
Department of Neurosurgery, University of Pittsburgh School of Medicine, 200 Lothrop St, Pittsburgh, PA, 15213, USA.
Hillman Comprehensive Cancer Center, University of Pittsburgh Medical Center, 5115 Centre Ave, Pittsburgh, PA, 15232, USA.
bioRxiv. 2023 May 31:2023.05.29.542774. doi: 10.1101/2023.05.29.542774.
Metabolism within the tumor microenvironment (TME) represents an increasing area of interest to understand glioma initiation and progression. Stable isotope tracing is a technique critical to the study of tumor metabolism. Cell culture models of this disease are not routinely cultured under physiologically relevant nutrient conditions and do not retain cellular heterogeneity present in the parental TME. Moreover, in vivo, stable isotope tracing in intracranial glioma xenografts, the gold standard for metabolic investigation, is time consuming and technically challenging. To provide insights into glioma metabolism in the presence of an intact TME, we performed stable isotope tracing analysis of patient-derived, heterocellular Surgically eXplanted Organoid (SXO) glioma models in human plasma-like medium (HPLM).
Glioma SXOs were established and cultured in conventional media or transitioned to HPLM. We evaluated SXO cytoarchitecture and histology, then performed spatial transcriptomic profiling to identify cellular populations and differential gene expression patterns. We performed stable isotope tracing with N-glutamine to evaluate intracellular metabolite labeling patterns.
Glioma SXOs cultured in HPLM retain cytoarchitecture and cellular constituents. Immune cells in HPLM-cultured SXOs demonstrated increased transcription of immune-related signatures, including innate immune, adaptive immune, and cytokine signaling programs. N isotope enrichment from glutamine was observed in metabolites from diverse pathways, and labeling patterns were stable over time.
To enable ex vivo, tractable investigations of whole tumor metabolism, we developed an approach to conduct stable isotope tracing in glioma SXOs cultured under physiologically relevant nutrient conditions. Under these conditions, SXOs maintained viability, composition, and metabolic activity while exhibiting increased immune-related transcriptional programs.
肿瘤微环境（TME）中的代谢是理解神经胶质瘤起始和进展的一个越来越受关注的领域。稳定同位素示踪是肿瘤代谢研究的一项关键技术。这种疾病的细胞培养模型通常不是在生理相关的营养条件下培养的，并且不能保留亲代TME中存在的细胞异质性。此外，在体内，颅内神经胶质瘤异种移植中的稳定同位素示踪是代谢研究的金标准，但耗时且技术上具有挑战性。为了深入了解完整TME存在下的神经胶质瘤代谢，我们在人血浆样培养基（HPLM）中对患者来源的异细胞手术切除类器官（SXO）神经胶质瘤模型进行了稳定同位素示踪分析。
建立神经胶质瘤SXO并在传统培养基中培养，或转换至HPLM中培养。我们评估了SXO的细胞结构和组织学，然后进行空间转录组分析以识别细胞群体和差异基因表达模式。我们用N-谷氨酰胺进行稳定同位素示踪以评估细胞内代谢物标记模式。
在HPLM中培养的神经胶质瘤SXO保留了细胞结构和细胞成分。HPLM培养的SXO中的免疫细胞显示出免疫相关特征的转录增加，包括先天免疫、适应性免疫和细胞因子信号传导程序。在来自不同途径的代谢物中观察到谷氨酰胺的N同位素富集，并且标记模式随时间稳定。
为了能够对整个肿瘤代谢进行体外易处理的研究，我们开发了一种方法，在生理相关营养条件下培养的神经胶质瘤SXO中进行稳定同位素示踪。在这些条件下，SXO保持活力、组成和代谢活性，同时表现出增加的免疫相关转录程序。
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