Dataset Information

GBM cell line secretome analysis

ABSTRACT: Glioblastoma (GBM) is the most common and aggressive brain tumor in adults. The purpose of the present study was to identify GBM cell-selective secreted proteins by analyzing conditioned media (CM) from GBM, breast, and colon cancer cell lines using sequential window acquisition of all theoretical spectra mass spectrometry (SWATH-MS) and targeted proteomics. We identified 26,041 peptides derived from 2,371 proteins in the CM from GBM and the other cancer cell lines. Among the proteins identified, 15 showed significantly higher expression in the CM from GBM cell lines than in those from other cancer cell lines.

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Sumio Ohtsuki

PROVIDER: PXD016898 | JPOST Repository | Sat Sep 19 00:00:00 BST 2020

REPOSITORIES: jPOST

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Publications

Laminin Subunit Alpha-4 and Osteopontin Are Glioblastoma-Selective Secreted Proteins That Are Increased in the Cerebrospinal Fluid of Glioblastoma Patients.

Kohata Tomohiro T Ito Shingo S Masuda Takeshi T Furuta Takuya T Nakada Mitsutoshi M Ohtsuki Sumio S

Journal of proteome research 20200716 8

Glioblastoma (GBM) is the most common and aggressive brain tumor in adults. The purpose of the present study was to identify GBM cell-selective secreted proteins by analyzing conditioned media (CM) from GBM, breast, and colon cancer cell lines using sequential window acquisition of all theoretical spectra-mass spectrometry (SWATH-MS) and targeted proteomics. We identified 2371 proteins in the CM from GBM and the other cancer cell lines. Among the proteins identified, 15 showed significantly high ...[more]

PMID: 32628487

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Project description:Background: Advanced Renal cell carcinoma (RCC) is therapeutically challenging. RCC progression is facilitated by mesenchymal stem/stromal cells (MSCs) that exert remarkable tumor tropism. The specific mechanisms mediating MSCs’ migration to RCC remain unknown. Here, we comprehensively analyzed RCC secretome to identify MSCs attractants. Methods: Conditioned media (CM) were collected from five RCC-derived cell lines (Caki-1, 786-O, A498, KIJ265T, KIJ308T) and non-tumorous control cell line (RPTEC/TERT1) and analyzed using cytokine arrays targeting 274 cytokines in addition to global CM proteomics. MSCs were isolated from bone marrow of patients undergoing standard orthopedic surgeries. RCC CM and the selected recombinant cytokines were used to analyze their influence on MSCs migration and microarray-targeted gene expression. The expression of genes encoding cytokines was evaluated in 100 matched-paired control-RCC tumor samples. Results: When compared with normal cells, CM from advanced RCC cell lines (Caki-1, KIJ265T) were the strongest stimulators of MSCs migration. Targeted analysis of 274 cytokines and global proteomics of RCC CM revealed decreased DPP4 and EGF, as well as increased AREG, FN1, and MMP1, with consistently altered gene expression in RCC cell lines and tumors. AREG and FN1 stimulated, while DPP4 attenuated MSCs migration. RCC CM induced MSCs’ transcriptional reprogramming, stimulating the expression of CD44, PTX3, and RAB27B. RCC cells secreted hyaluronic acid (HA), a CD44 ligand mediating MSCs’ homing to the kidney. AREG emerged as an upregulator of MSCs’ transcription. Conclusions: advanced RCC cells secrete AREG, FN1 and HA to induce MSCs migration, while DPP4 loss prevents its inhibitory effect on MSCs homing. RCC secretome induces MSCs’ transcriptional reprograming to facilitate their migration. The identified components of RCC secretome represent potential therapeutic targets. We used microarrays to determine the effect of the conditioned media (CM) collected from two RCC-derived cell lines (Caki-1, KIJ265T) and non-tumorous control cell line (RPTEC/TERT1) on the transcriptome change in mesenchymal stem/stromal cells (MSCs).

Project description:Background: Radiation therapy is the standard of care for central nervous system tumours. Despite the success of radiation therapy in reducing tumour mass, irradiation (IR)-induced vasculopathies and neuroinflammation contribute to late-delayed complications, neurodegeneration, andpremature ageing in long-term cancer survivors. Mesenchymal stromal cells (MSCs) are adult stem cells that facilitate tissue integrity, homeostasis, and repair. Here, we investigated the potential of the iPSC-derived MSC(iMSC) secretome in immunomodulation and vasculature repair in response to radiation injury utilizing human cell lines. Methods: We generated iPSC-derived iMSC lines and evaluated the potential of their conditioned media (iMSC CM) to treat IR-induced injuries in human monocytes (THP1) and brain vascular endothelial cells (hCMEC/D3). We further assessed factors in the iMSC secretome, their modulation, and the molecular pathways they elicit. Results: Increasing doses of IR disturbed endothelial tube and spheroid formation in hCMEC/D3. When IR-injured hCMEC/D3 (IR ≤ 5 Gy) were treated with iMSC CM, endothelial cell viability, adherence, spheroid compactness, and proangiogenic sprout formation were significantly ameliorated, and IR-induced ROS levels were reduced. iMSC CM augmented tube formation in cocultures of hCMEC/D3 and iMSCs. Consistently, iMSC CM facilitated angiogenesis in a zebrafish model in vivo. Furthermore, iMSC CM suppressed IR-induced NFκB activation, TNF-α release, and ROS production in THP1 cells. Additionally, iMSC CM diminished NF-kB activation in THP1 cells cocultured with irradiated hCMEC/D3, iMSCs, or HMC3 microglial lines. The cytokine array revealed that iMSC CM contains the proangiogenic and immunosuppressive factors MCP1/CCL2, IL6, IL8/CXCL8, ANG (Angiogenin), GROα/CXCL1, and RANTES/CCL5. Common promoter regulatory elements were enriched in TF-binding motifs such as androgen receptor (ANDR) and GATA2. hCMEC/D3 phosphokinome profiling revealed increased expression of pro-survival factors, the PI3K/AKT/mTOR modulator PRAS40 and β-catenin in response to CM. The transcriptome analysis revealed increased expression of GATA2 in iMSCs and the enrichment of pathways involved in RNA metabolism, translation, mitochondrial respiration, DNA damage repair, and neurodevelopment. Conclusions: The iMSC secretome is a comodulated composite of proangiogenic and immunosuppressive factors that has the potential to alleviate radiation-induced vascular endothelial cell damage and immune activation.

Project description:Rationale: In virtually all models of heart failure, prognosis is determined by right ventricular (RV) function; thus, understanding the cellular mechanisms contributing to RV dysfunction is critical. Whole organ remodeling is associated with cell-specific changes, including cardiomyocyte dedifferentiation and activation of cardiac fibroblasts (Cfib) which in turn is linked to disorganization of cytoskeletal proteins and loss of sarcomeric structures. However, how these cellular changes contribute to RV function remains unknown. We’ve previously shown significant organ-level RV dysfunction in a large animal model of pulmonary hypertension (PH) which was not mirrored by reduced function of isolated cardiomyocytes. We hypothesized that factors produced by the endogenous Cfib contribute to global RV dysfunction by generating a heterogeneous cellular environment populated by dedifferentiated cells. Objective: To determine the effect of Cfib conditioned media (CM) from the PH calf (PH-CM) on adult rat ventricular myocytes (ARVM) in culture. Methods and Results: Brief exposure (<2 days) to PH-CM results in rapid, marked dedifferentiation of ARVM to a neonatal-like phenotype exhibiting spontaneous contractile behavior. Dedifferentiated cells maintain viability for over 30 days with continued expression of cardiomyocyte proteins including TnI and α-actinin yet exhibit myofibroblast characteristics including expression of α-smooth muscle actin. Using a bioinformatics approach to identify factor(s) that contribute to dedifferentiation, we found activation of the PH Cfib results in a unique transcriptome correlating with factors both in the secretome and with activated pathways in the dedifferentiated myocyte. Further, we identified upregulation of periostin in the Cfib and CM, and demonstrate that periostin is sufficient to drive cardiomyocyte dedifferentiation. Conclusions: These data suggest that paracrine factor(s) released by Cfib from the PH calf signal a phenotypic transformation in a population of cardiomyocytes that likely contributes to RV dysfunction. Therapies targeting this process, such as inhibition of periostin, have the potential to prevent RV dysfunction.

			Action	DRS
	Caco-2_Mix-24h.wiff	Wiff
	Caco-2_Mix-24h.wiff.scan	Wiff
	Caco-2_Mix_48h.wiff	Wiff
	Caco-2_Mix_48h.wiff.scan	Wiff
	HEPG2_Mix_24h.wiff	Wiff