Decoding the role of macrophages in periodontitis and type 2 diabetes using single‐cell RNA‐sequencing

2022 ◽  
Vol 36 (2) ◽  
Author(s):  
Panagiota Agrafioti ◽  
Joshua Morin‐Baxter ◽  
Kranthi K. K. Tanagala ◽  
Sunil Dubey ◽  
Peter Sims ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Single Cell ◽  
Rna Sequencing ◽  
Single Cell Rna Sequencing

scholarly journals A Transcriptomic and Proteomic Atlas of Obesity and Type 2 Diabetes in Cynomolgus Monkeys

2021 ◽  
Author(s):  
Xianglong Zhang ◽  
Ying Lei ◽  
Oliver Homann ◽  
Marina Stolina ◽  
Songli Wang ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
Immune Response ◽  
Nonhuman Primate ◽  
Metabolic Disorders ◽  
Healthy Controls ◽  
Adipose Tissues ◽  
Cynomolgus Monkeys ◽  
Single Cell Rna Sequencing

Obesity and type 2 diabetes (T2D) remain major global healthcare challenges and developing therapeutics necessitate using nonhuman primate models. Here, we present transcriptomic and proteomic analyses of all the major organs of cynomolgus monkeys with spontaneous obesity or T2D in comparison to healthy controls. Molecular changes occur predominantly in the adipose tissues of individuals with obesity, while extensive expression perturbations among T2D individuals are observed in many tissues, such as the liver, kidney, brain, and heart. Immune response-related pathways are upregulated in obesity and T2D, whereas metabolism and mitochondrial pathways are downregulated. Incorporating human single-cell RNA sequencing findings corroborates the role of macrophages and monocytes in obesity. Moreover, we highlight some potential therapeutic targets including SLC2A1 and PCSK1 in obesity as well as SLC30A8 and SLC2A2 in T2D. Our findings provide insights into tissue-specific molecular foundations of obesity and T2D and reveal the mechanistic links between these two metabolic disorders.

Exploring the role of the skin microenvironment in cutaneous T-cell lymphoma using single cell RNA-sequencing

2021 ◽  
Vol 156 ◽  
pp. S3-S4
Author(s):  
Gabor Dobos ◽  
Andreea Calugareanu ◽  
Laurence Michel ◽  
Maxime Battistella ◽  
Caroline Ram-Wolff ◽  
...  
Keyword(s):  
T Cell ◽  
Single Cell ◽  
Rna Sequencing ◽  
Cell Lymphoma ◽  
T Cell Lymphoma ◽  
Cutaneous T Cell Lymphoma ◽  
Single Cell Rna Sequencing

scholarly journals Weighted Gene Coexpression Network Analysis Identified MicroRNA Coexpression Modules and Related Pathways in Type 2 Diabetes Mellitus

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
Tianyu Feng ◽  
Kexin Li ◽  
Pingping Zheng ◽  
Yanjun Wang ◽  
Yaogai Lv ◽  
...  
Keyword(s):  
Diabetes Mellitus ◽  
Type 2 Diabetes ◽  
Type 2 Diabetes Mellitus ◽  
Network Analysis ◽  
Rna Sequencing ◽  
Gene Coexpression Network ◽  
Gene Coexpression ◽  
Coexpression Network Analysis

Objective. Type 2 diabetes mellitus (T2DM) is a metabolic disease with high incidence, which has seriously affected human life and health. MicroRNA, a short-chain noncoding RNA, plays an important role in T2DM. Identification of meaningful microRNA modules and the role of microRNAs provide a basis for searching potential biomarkers of T2DM. Materials and Methods. In this study, three newly diagnosed patients with T2DM and three controls were selected for Whole Peripheral Blood RNA Sequencing to establish a microRNA library. Weighted gene coexpression network analysis (WGCNA) was applied to construct coexpression modules and to detect the trait-related microRNA modules; then, KEGG enrichment analysis was performed to predict the biological function of the interest modules, and candidate hub microRNAs were screened out by the value of module membership (MM) and protein-protein interaction (PPI) network. Result. Four microRNA modules (blue, brown, magenta, and turquoise) were highly associated with the T2DM; the number of miRNAs in these modules ranged from 41 to 469. The Fc gamma R-mediated phagocytosis pathway, Rap1 signaling pathway, MAPK signaling pathway, and Lysosome pathway were common pathways in three of the four modules. RPS27A, UBC, and RAC1 were the top three proteins in our study; their corresponding RNAs were miR-1271-5p, miR-130a-3p, miR-130b-3p, and miR-574-3p. Conclusion. In summary, this study identified blood miRNAs in human T2DM using RNA sequencing. The findings may be the foundation for understanding the potential role of miRNAs in T2DM.

scholarly journals Human Visceral Adipose Tissue (VAT) Inflammation Is Not Associated With Perforin Deficiency in Type 2 Diabetes Mellitus (DM) Subjects as Compared to Health Controls

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. A15-A16
Author(s):  
Ravindra Kumar Shukla ◽  
Ramkaran Choudhary ◽  
Mahendra Lodha ◽  
Mahaveer singh Rodha ◽  
Nitin Bajpai ◽  
...  
Keyword(s):  
Type 2 Diabetes ◽  
T Cell ◽  
Nk Cells ◽  
Single Cell ◽  
Stromal Vascular Fraction ◽  
Healthy Controls ◽  
Type 2 Dm ◽  
And Control

Abstract Introduction: Role of T cells in VAT inflammation is poorly characterised. Perforin deficiency is associated with autoimmune inflammatory diseases like Hemophagocytosis Lymphohistiocytosis,as also in context of type 1 DM pathogenesis. Data from animal models suggest that perforin deficiency leads to VAT inflammation1Objective: We hypothesized CD8+/perf+ and CD56+/perf+ cells to be decreased in type 2 DM as compared to healthy controls. The present study also explored the difference in activation of T/NK cells between two groups Methods: 2×2 cm omental tissue was obtained from subjects undergoing elective abdominal surgery. The sample was transported in RPMI solution and stored in -80 ℃. Processing involved thawing, incubation at 37.6 ℃ for 24 hours with type IV Collagenase (1 mg/ml, Sigma Aldrich) 1ml/g of tissue,centrifuge (32g for 10 min at 10℃). The resultant Stromal Vascular fraction (SVF) was suspended in phosphate buffer saline (PBS), passed through cell strainer to make single cell suspension. It was again centrifuged and tagged with CD markers of interest. Fc block was added and single cell solution with FACS fluid prepared. It was run in BD CANTO-2 flow cytometer as described2Results: Of seventeen samples analysed, twelve samples of type 2 diabetes subjects were compared with five healthy controls. All results are presented in median. The diabetics had higher HbA1c (8.1 % vs 6%), higher BMI (28 kg/m2 vs 24 kg/m2), hsCRP(2.1 mg/dl Vs 0.9 mg/dl) but there was no difference in HOMA-Ir (5 vs 5.2 mU/L/mg/dl). The percentage of CD4+ + CD8+ cells/g of VAT was similar in both cases and control (20 × 103 Vs 23× 103). CD8+/CD45+/perf+ and CD56+/CD45+/perf+ could not be identified in any of the samples. Although CD8+/CD45+/perf-- and CD56+/CD45+/perf-- cells were identified. Next, we analysed the same cells for cytotoxic activation by 107a. The percentage of 107a positivity was low in CD 8+ (7% and 4 % respectively in cases and control) and CD 56+ cells (10% and 9 % respectively in cases and control),Although clinically type 2 DM subjects were obese and had inflammation (i.e higher hsCRP), there was no difference in VAT activation of immune cells studied. Also, we could not delineate perforin in any of the samples. Conclusion: Taken together this work suggests VAT T cell immune milieu in human Type 2 DM is different from mouse model. It is neither characterised by perforin deficiency nor activation of T cell/NK cell. This study points towards the probability that, the role of T cell/NK cells in human VAT infiltration could be fundamentally different from mice models. Further studies should be focussed on functional characteristics of these cells and interaction with VAT macrophages. References 1. Xavier S. Revelo et al Diabetes 2015;64:90–103 2. Wetzels S et al J Vis Exp. 2018 Mar 6;(133):57319.

scholarly journals EPCO-32. AN EPIGENETIC SINGLE-CELL ATLAS OF IDH-MUTANT GLIOMA REVEALS THE ROLE OF ATRX IN SHAPING TUMOR COMPOSITION

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii76-ii76
Author(s):  
Husam Babikir ◽  
Lin Wang ◽  
Karin Shamardani ◽  
Sweta Sudhir ◽  
Gary Kohanbash ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Single Cell ◽  
Rna Sequencing ◽  
Myeloid Cell ◽  
Cell Types ◽  
Open Chromatin ◽  
Helix Loop Helix ◽  
Local Immunosuppression ◽  
Single Cell Rna Sequencing

Abstract Recent single-cell RNA-sequencing studies have identified a hierarchy of cell types that is common to all isocitrate dehydrogenase (IDH) -mutant gliomas. This finding is somewhat paradoxical since the genetic differences between IDH-mutant astrocytomas and IDH-mutant oligodendrogliomas are prognostic, predictive of therapeutic response, and correlated with differences in immune infiltrates. To integrate these disparate findings, we constructed a single-cell atlas of 28 human IDH-mutant primary untreated grade-II/III gliomas. All specimens were profiled by single-cell assay for transposase-accessible chromatin, with additional cohorts profiled via single-cell RNA-sequencing and single-cell spatial proteomics. We determined the cell-type specific differences between IDH-mutant gliomas in transcription-factor utilization, associated targeting and cis-regulatory grammars. To elucidate the role of the chromatin remodeler ATRX (inactivated in over 86% of IDH-mutant astrocytomas) in shaping observed differences in open chromatin, we knocked out ATRX in an immunocompetent model of IDH-mutant glioma and subjected murine tumors to single-cell profiling. We found: 1. ATRX-deficient, IDH-mutant human and murine gliomas both upregulate an astrocytic regulatory program driven by Nuclear Factor I genes and downregulate an oligodendrocytic program driven by basic helix-loop-helix transcription factors. 2. Both human and mouse ATRX-deficient, IDH-mutant gliomas up-regulate genes that promote myeloid-cell chemotaxis and both have significantly higher percentages of myeloid-derived immune-suppressive cells than controls; 3. A transcription-factor program is conserved between human and murine ATRX-deficient tumors that shapes glial identity and promotes local immunosuppression. These studies elucidate how IDH-mutant gliomas from different subtypes can have distinct cellular morphologies and tumor micronenvironments despite a common lineage hierarchy.

scholarly journals Calcium-Prolactin Secretion Coupling in Rat Pituitary Lactotrophs Is Controlled by PI4-Kinase Alpha

2021 ◽  
Vol 12 ◽  
Author(s):  
Marek Kučka ◽  
Arturo E. Gonzalez-Iglesias ◽  
Melanija Tomić ◽  
Rafael M. Prévide ◽  
Kosara Smiljanic ◽  
...  
Keyword(s):  
Calcium Signaling ◽  
Single Cell ◽  
Rna Sequencing ◽  
Intracellular Signaling ◽  
Specific Inhibitor ◽  
Pcr Analysis ◽  
Single Cell Rna Sequencing ◽  
Specific Inhibitors ◽  
Calcium Secretion

The role of calcium, but not of other intracellular signaling molecules, in the release of pituitary hormones by exocytosis is well established. Here, we analyzed the contribution of phosphatidylinositol kinases (PIKs) to calcium-driven prolactin (PRL) release in pituitary lactotrophs: PI4Ks - which control PI4P production, PIP5Ks - which synthesize PI(4, 5)P2 by phosphorylating the D-5 position of the inositol ring of PI4P, and PI3KCs – which phosphorylate PI(4, 5)P2 to generate PI(3, 4, 5)P3. We used common and PIK-specific inhibitors to evaluate the strength of calcium-secretion coupling in rat lactotrophs. Gene expression was analyzed by single-cell RNA sequencing and qRT-PCR analysis; intracellular and released hormones were assessed by radioimmunoassay and ELISA; and single-cell calcium signaling was recorded by Fura 2 imaging. Single-cell RNA sequencing revealed the expression of Pi4ka, Pi4kb, Pi4k2a, Pi4k2b, Pip5k1a, Pip5k1c, and Pik3ca, as well as Pikfyve and Pip4k2c, in lactotrophs. Wortmannin, a PI3K and PI4K inhibitor, but not LY294002, a PI3K inhibitor, blocked spontaneous action potential driven PRL release with a half-time of ~20 min when applied in 10 µM concentration, leading to accumulation of intracellular PRL content. Wortmannin also inhibited increase in PRL release by high potassium, the calcium channel agonist Bay K8644, and calcium mobilizing thyrotropin-releasing hormone without affecting accompanying calcium signaling. GSK-A1, a specific inhibitor of PI4KA, also inhibited calcium-driven PRL secretion without affecting calcium signaling and Prl expression. In contrast, PIK93, a specific inhibitor of PI4KB, and ISA2011B and UNC3230, specific inhibitors of PIP5K1A and PIP5K1C, respectively, did not affect PRL release. These experiments revealed a key role of PI4KA in calcium-secretion coupling in pituitary lactotrophs downstream of voltage-gated and PI(4, 5)P2-dependent calcium signaling.

scholarly journals Single-Cell RNA Sequencing in Multiple Pathologic Types of Renal Cell Carcinoma Revealed Novel Potential Tumor-Specific Markers

2021 ◽  
Vol 11 ◽  
Author(s):  
Cheng Su ◽  
Yufang Lv ◽  
Wenhao Lu ◽  
Zhenyuan Yu ◽  
Yu Ye ◽  
...  
Keyword(s):  
Renal Cell Carcinoma ◽  
Cell Carcinoma ◽  
Single Cell ◽  
Rna Sequencing ◽  
Renal Cell ◽  
Small Sample Size ◽  
Mrna Level ◽  
Small Sample ◽  
Single Cell Rna Sequencing

BackgroundRenal cell carcinoma (RCC) is the most common type of kidney cancer. Studying the pathogenesis of RCC is particularly important, because it could provide a direct guide for clinical treatment. Given that tumor heterogeneity is probably reflected at the mRNA level, the study of mRNA in RCC may reveal some potential tumor-specific markers, especially single-cell RNA sequencing (scRNA-seq).MethodsWe performed an exploratory study on three pathological types of RCC with a small sample size. This study presented clear-cell RCC (ccRCC), type 2 pRCC, and chRCC in a total of 30,263 high-quality single-cell transcriptome information from three pathological types of RCC. In addition, scRNA-seq was performed on normal kidneys. Tumor characteristics were well identified by the comparison between different pathological types of RCC and normal kidneys at the scRNA level.ResultsSome new tumor-specific markers for different pathologic types of RCC, such as SPOCK1, PTGIS, REG1A, CP and SPAG4 were identified and validated. We also discovered that NDUFA4L2 both highly expressed in tumor cells of ccRCC and type 2 pRCC. The presence of two different types of endothelial cells in ccRCC and type 2 pRCC was also identified and verified. An endothelial cell in ccRCC may be associated with fibroblasts and significantly expressed fibroblast markers, such as POSTN and COL3A1. At last, by applying scRNA-seq results, the activation of drug target pathways and sensitivity to drug responses was predicted in different pathological types of RCC.ConclusionsTaken together, these findings considerably enriched the single-cell transcriptomic information for RCC, thereby providing new insights into the diagnosis and treatment of RCC.

scholarly journals TMIC-06. MYELOID POPULATIONS AND THE EFFECT OF NEOADJUVANT PD-1 INHIBITION IN THE GLIOBLASTOMA MICROENVIRONMENT: A SURFACEOMIC AND TRANSCRIPTOMIC DISSECTION AT THE SINGLE-CELL LEVEL

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi248-vi248
Author(s):  
Aaron Mochizuki ◽  
Alexander Lee ◽  
Joey Orpilla ◽  
Jenny Kienzler ◽  
Mildred Galvez ◽  
...  
Keyword(s):  
Brain Tumor ◽  
T Cell ◽  
Single Cell ◽  
Rna Sequencing ◽  
Myeloid Cells ◽  
Single Cell Level ◽  
Mass Cytometry ◽  
Cell Level ◽  
Single Cell Rna Sequencing

Abstract INTRODUCTION Glioblastoma (GBM) is the most common malignant brain tumor in adults and is associated with a dismal prognosis. Neoadjuvant anti-PD-1 blockade has demonstrated efficacy in melanoma, non-small cell lung cancer and recurrent GBM; however, responses vary. While T cells have garnered considerable attention in the context of immunotherapy, the role of myeloid cells in the GBM microenvironment remains controversial. METHODS We isolated CD45+ immune populations from patients who underwent brain tumor resection at UCLA. We hypothesized that myeloid cells in glioblastoma contribute to T cell dysfunction; however, this immune suppression can be mitigated by neoadjuvant PD-1 inhibition. To test this, we utilized mass cytometry and single-cell RNA sequencing to characterize these immune populations. RESULTS Mass cytometry profiling of tumor infiltrating lymphocytes from patients with GBM demonstrated a preponderance of CD11b+ myeloid populations (75% versus 25% CD3+). At the transcriptomic level, myeloid cells in newly diagnosed GBMs exhibited decreased expression of CCL4 (loge fold change -1.18, Bonferroni-adjusted P = 1.62x10-254) and its ligands compared to anaplastic astrocytoma. In ranked gene set enrichment analysis, patients who received neoadjuvant pembrolizumab demonstrated enrichment in TNFα-, NFκB- and lipid metabolism-related gene sets by bootstrapped Kolmogorov-Smirnov test (Benjamini-Hochberg adjusted P = 4.74x10-3, 1.45x10-2 and 2.48x10-3, respectively) in tumor-associated myeloid populations. Additionally, single-cell trajectory analysis demonstrated increased CCL4 and decreased ISG15 with neoadjuvant checkpoint inhibition. CONCLUSIONS Here, we utilize mass cytometry and single-cell RNA sequencing to demonstrate the predominance and transcriptomic features of myeloid populations in GBM. Myeloid cells in patients who receive neoadjuvant PD-1 blockade re-express increased levels NFκB, TNFα and CCL4, a cytokine crucial for the recruitment of dendritic cells to the tumor for antigen-specific T cell activation. By delving into the GBM microenvironment at the single-cell level, we hope to better delineate the role of myeloid populations in this uniformly fatal tumor.

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