Project description:To uncover the natural cell heterogeneity state of healthy eccrine sweat gland (ESG) at the gene level, we obtained normal human skin tissue from three subjects without clinical evidence or a history of inflammatory or systemic skin disease and isolated the ESG performed single-cell (sc) RNA sequencing assay.
Project description:The eccrine sweat gland is an exocrine gland that is involved in the secretion of sweat for control of temperature. Malfunction of the sweat glands can result in disorders such as miliaria, hyperhidrosis and bromhidrosis. In addition, inadequate reabsorption of salt from sweat is a major feature of cystic fibrosis. Understanding the transcriptome and proteome of sweat glands is important for understanding the physiology and the role in disease. However, no systematic transcriptome or proteome analysis of sweat glands has yet been reported. To this end, we isolated eccrine sweat glands by microdissecting them from human skin and performed both RNA-seq and proteome analysis. In total, ~138,000 transcripts and ~6,100 proteins were identified. The proteome data of eccrine sweat gland showed enrichment of proteins involved in secretion, reabsorption, and wound healing while the transcriptome data did not show any enrichment for a specific pathway. Importantly, protein level identification of TRPV4 in eccrine sweat gland establishes its importance in re-epithelialization of partial-thickness wound and prevention of dehydration. Furthermore, this study enabled us to identify2 missing proteins. Integration of RNA-seq and proteomic data allowed us to identify 7 peptides from 5 novel genes. Most of the novel proteins were from short open reading frames (sORFs) suggesting that many sORFs still remain to be annotated in the human genome. The peptides mapping to the missing or novel proteins were validated by analyzing synthetic peptides. This study provides the first integrated analysis of the transcriptome and proteome of the human eccrine sweat gland and should become an invaluable resource to biomedical research community for studying sweat glands in physiology and disease.
Project description:Adult human fibroblasts derived from the dermis of 4 mm punch biopsies taken from the lower backs of 15 healthy subjects of 3 different phototypes (types I, III and VI). This study was approved by the Human Research Ethics Committee of Hamburg. Subjects with type I skin were #4, 12, 15, 19 and 26; subjects with type III skin were # 6, 9, 13, 14 and 23, and subjects with type VI skin were # 7, 8, 16, 17 and 28. The fibroblasts were cultured from the biopsies and were grown in monolayer culture in high glucose Dulbecco’s modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and 1% L-glutamine plus 1% penicillin and streptomycin at 37°C in a humidified 5% CO2 atmosphere. Fibroblasts were subcultured using routine methods and were used at passages 3 to 6.
Project description:The eccrine sweat gland is an exocrine gland that is involved in the secretion of sweat for control of temperature. Malfunction of the sweat glands can result in disorders such as miliaria, hyperhidrosis and bromhidrosis. In addition, inadequate reabsorption of salt from sweat is a major feature of cystic fibrosis. Understanding the transcriptome and proteome of sweat glands is important for understanding the physiology and the role in disease. However, no systematic transcriptome or proteome analysis of sweat glands has yet been reported. To this end, we isolated eccrine sweat glands by microdissecting them from human skin and performed both RNA-seq and proteome analysis. In total, ~138,000 transcripts and ~6,100 proteins were identified. The proteome data of eccrine sweat gland showed enrichment of proteins involved in secretion, reabsorption, and wound healing while the transcriptome data did not show any enrichment for a specific pathway. Importantly, protein level identification of TRPV4 in eccrine sweat gland establishes its importance in re-epithelialization of partial-thickness wound and prevention of dehydration. Furthermore, this study enabled us to identify2 missing proteins. Integration of RNA-seq and proteomic data allowed us to identify 7 peptides from 5 novel genes. Most of the novel proteins were from short open reading frames (sORFs) suggesting that many sORFs still remain to be annotated in the human genome. The peptides mapping to the missing or novel proteins were validated by analyzing synthetic peptides. This study provides the first integrated analysis of the transcriptome and proteome of the human eccrine sweat gland and should become an invaluable resource to biomedical research community for studying sweat glands in physiology and disease.
Project description:Diabetic foot ulcers (DFUs) are a devastating complication of diabetes. In order to identify systemic and local factors associated with DFU healing, we examined the cellular landscape of DFUs by single-cell RNA-seq analysis of foot and forearm skin specimens, as well as PBMC samples, from 10 non-diabetic subjects, and 17 diabetic patients, 11 with, and 6 without DFU. Our analysis shows enrichment of a unique inflammatory fibroblast population in DFU patients with healing wounds. The patients with healing DFUs also depicted enrichment of macrophages with M1 polarization, as opposed to more M2 macrophages in non-healing wounds. These findings were verified using Immunohistochemistry and Spatial Transcriptomics.
Project description:We performed RNA-seq to analyze gene expression in human PASMCs (Pulmonary arterial smooth muscle cells) isolated from subjects without disease and from subjects with IPAH (idiopathic pulmonary hypertension)
Project description:Purpose: The goal of this study to use ethanol-exposed human embryonic stem cell (hESC)-derived neural cells as models to investigate mRNA expression changes in the brains of subjects with alcohol use disorder (AUD). Methods: hESCs were differentiated into neural cells (mainly cortical interneurons), which were then cultured in media with or without ethanol (50-100 mM) for 7 days (by duplicate experiments). Total RNAs were extracted from hESC-derived neural cells (with or without ethanol exposure) for mRNA sequencing. The sequence reads were processed using the RNA-seq processing pipeline Pipeliner [Front Genet. 2019; 10:614] workflow. Ethanol-induced mRNA transcriptomic changes were analyzed by the Limma-Voom method. Results: When the significance level was set at FC>2.0 & P<0.01, 19 coding genes showed differential expression, and all of them were downregulated after a 7-day ethanol exposure. Conclusions: The hESC-derived neural cell model study can partially validate mRNA transcriptomic changes in postmortem brains of subjects with alcohol use disorder.
Project description:We established a culture method of human keratinocytes from the bulge region of a plucked hair follicle, that contains multipotent epithelial stem cells with high proliferative potential. Using our method, keratinocyte cultures were successfully obtained from all subjects without invasive skin biopsies. We compared the gene expression profiles between the cultured keratinocytes derived from human hair-follicle-bulge (bulge–derived keratinocytes; BDKs) and neonatal human epidermal keratinocytes (NHEKs), and between BDKs from donors with atopic dermatitis and non-atopic controls using microarray analysis. Keywords: expressin profiling
Project description:In order to better understand the molecular mechanisms underlying right ventricular failure in human PAH, a RNA sequencing analysis was performed in RV tissues obtained from subjects clinically categorized as compensated RV (n=11), decompensated RV (n=7) and donor controls (n=14). The study unveils a plethora of factors dysregulated in the RV of PAH patients, which opens new perspectives for translational research.