nuclei-isolation-from-fresh-frozen-brain-tumors-for-single-nucleus

Nuclei Isolation from Fresh Frozen Brain Tumors for Single-Nucleus RNA-seq and ATAC-seq

Adult diffuse gliomas exhibit inter- and intra-tumor heterogeneity. Until recently, the majority of large-scale molecular profiling efforts have focused ...

University of Heidelberg, INF 400, Heidelberg, Germany

Vascular endothelial growth factor (VEGF) is one of the key factors in tumor neoangiogenesis, acting through its receptors KDR (VEGFR-2) and fit-1 (VEGFR-1) expressed on endothelial cells. Our data demonstrate that VEGFR-1 and to a lesser extent VEGFR-2 are expressed in a number of human tumor tissues and derived cells in culture. VEGFR-1 protein is expressed in 26 of 42 glioma tissues, 22 of which show a coexpression of VEGFR-1 with VEGFR-2; 1 glioma tissue expresses exclusively VEGFR-2. In the derived glioma cell cultures, we found VEGFR-1 mRNA expression in 6 of 11 cultures, with one coexpressing VEGFR-1 and VEGFR-2. Of four established glioma cell lines, two expressed VEGFR-1. In addition VEGFR-1 protein expression was demonstrated in 30 of 37 tumor tissues of squamous cell carcinomas of the head and neck, with VEGFR-2 coexpression in 15 tissues and an expression of VEGFR-2 alone in 1 tissue. Derived tumor cell cultures showed mRNA expression of VEGFR-1 alone in seven of seven cases. Established melanoma cell lines expressed VEGFR-1 mRNA in four of five lines, with VEGFR-2 coexpression in two lines. Concerning the functional significance of VEGF receptor expression, VEGF treatment of VEGFR-1-expressing tumor cells induced the inhibition of cell proliferation by 25 to 55% and the inhibition of tumor cell migration by 29 to 55%. Thus our data indicate that the coexpression of VEGF and VEGFR-1 in tumor cells could have an inhibitory effect on tumor cell proliferation and migration, a mechanism possibly induced as a response to a deficiency in nutrient and oxygen supply.

Expression and functional significance of vascular endothelial growth factor receptors in human tumor cells.

The extracellular matrix protein tenascin-C is expressed in processes like embryogenesis and wound healing and in neoplasia. Tenascin-C expression in gliomas has been described previously; however, the relation to clinical data remains inconsistent. Generally, analysis of tenascin-C function is difficult due to different alternatively spliced isoforms. Our studies focus on changes in tenascin-C expression in human gliomas, correlating these changes with tumor progression and elucidating the functional role of the glioma cell-specific tenascin-C isoform pool. Eighty-six glioma tissues of different World Health Organization (WHO) grades were analyzed immunohistochemically for tenascin-C expression. The influence of the specific tenascin-C isoforms produced by glioblastoma cells on proliferation and migration was examined in vitro using blocking antibodies recognizing all isoforms. In general, tenascin-C expression increased with tumor malignancy. Perivascular staining of tenascin-C around tumor-supplying blood vessels was observed in all glioblastoma tissues, whereas in WHO II and III gliomas, perivascular tenascin-C staining appeared less frequently. The appearance of perivascular tenascin-C correlated significantly with a shorter disease-free time. Analysis of proliferation and migration in the presence of blocking antibodies revealed an inhibition of proliferation by around 30% in all 3 glioblastoma cell cultures, as well as a decrease in migration of 30.6-46.7%. Thus we conclude that the endogenous pool of tenascin-C isoforms in gliomas supports both tumor cell proliferation and tumor cell migration. In addition, our data on the perivascular staining of tenascin-C in WHO II and III gliomas and its correlation with a shorter disease-free time suggest that tenascin-C may be a new and potent prognostic marker for an earlier tumor recurrence.

Clinical impact and functional aspects of tenascin-C expression during glioma progression.

A major part of non-small cell lung cancer (NSCLC) patients treated with mono- or multimodal concept develop therapy resistance. Despite the abundance of biomarkers investigated in the past, there is still a need for valid NSCLC biomarkers. Glycodelin, an immunosuppressive endometrial protein, has been shown to be also expressed in NSCLC. Here, we investigated its potential as a biomarker in metastatic and advanced stage NSCLC. Glycodelin gene and protein expression were measured in 28 therapy-naïve resected tumors as well as in corresponding brain ( = 16) and adrenal gland ( = 12) metastasis by qPCR and IHC. Moreover, we correlated glycodelin gene expression of cryoconserved therapy-naïve biopsies ( = 55) of advanced stage patients with glycodelin serum concentrations and patient survival. Using follow-up samples of the patients, we monitored glycodelin serum concentrations during therapy. Glycodelin expression correlated between primary tumor and distant metastases within the same patients. The gene expression of glycodelin in therapy-naïve biopsies also correlated with the serum concentrations of the patients (r = 0.60). Patients with elevated serum concentrations showed a tendency in lower overall survival ( = 0.088) and measuring of glycodelin indicated a progression of the disease earlier compared to clinical diagnostic. Taken together, we demonstrate that glycodelin is a promising prognostic and follow-up biomarker for metastatic and advanced NSCLC.

Glycodelin as a Serum and Tissue Biomarker for Metastatic and Advanced NSCLC.

Translational cancer research has seen an increasing interest in metabolomic profiling to decipher tumor phenotypes. However, the impact of post-surgical freezing delays on mass spectrometric metabolomic measurements of the cancer tissue remains elusive.

Article	Year
Expression and functional significance of vascular endothelial growth factor receptors in human tumor cells. Laboratory investigation; a journal of technical methods and pathology\| PubMed ID: 10616207	1999
Clinical impact and functional aspects of tenascin-C expression during glioma progression. International journal of cancer\| PubMed ID: 11920587	2002
Glycodelin as a Serum and Tissue Biomarker for Metastatic and Advanced NSCLC. Cancers\| PubMed ID: 30518088	2018
Impact of post-surgical freezing delay on brain tumor metabolomics. Metabolomics : Official journal of the Metabolomic Society\| PubMed ID: 31087206	2019

Christel Herold-Mende

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