scholarly journals Cross-tissue single-cell transcriptomic landscape reveals the key cell subtypes and their potential roles in the nutrient absorption and metabolism in dairy cattle

2021 ◽  
Author(s):  
Jia-Jin Wu ◽  
Senlin Zhu ◽  
Fengfei Gu ◽  
Teresa G. Valencak ◽  
Jian-Xin Liu ◽  
...  
Keyword(s):  
Dairy Cattle ◽  
Single Cell ◽  
Nutrient Absorption

scholarly journals Single-cell RNA-Aequencing Reveals Novel Myofibroblasts with Epithelial Cell-Like Features in the Mammary Gland of Dairy Cattle

2020 ◽  
Author(s):  
Fengfei Gu ◽  
Jiajin Wu ◽  
Senlin Zhu ◽  
Teresa G. Valencak ◽  
Jian-Xin Liu ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Dairy Cattle ◽  
Single Cell ◽  
Dairy Product ◽  
Cell Types ◽  
Marker Genes ◽  
New Findings ◽  
Luminal Cells

Abstract Background: Cow’s milk is a highly-nutritious dairy product that is widely consumed worldwide. It is secreted by the developed mammary gland (MG) of dairy cattle. However, a comprehensive understanding of cell-type diversity and cell function within bovine MG is lacking. In the current study, we used single-cell RNA sequencing to investigate the transcriptome of 24,472 high-quality MG cells isolated from newborn and adult cows. Results: Unbiased clustering analysis revealed the existence of 24 cell types, which could be divided into four categories: 9 immune, 3 epithelial, 9 fibroblast, and 3 endothelial cell types. Other cell subtypes were further identified based on re-clustering and pseudotemporal reconstruction of epithelial cells that included 3 mature luminal epithelial, 1 intermediate, and 2 progenitor cell subtypes. The individual top marker genes of these 3 mature luminal epithelial cell subtypes (L0, L1, and L5) were APOA1, STC2, and PTX3, which were further validated using immunofluorescence. Based on functional analysis, the L0, L1, and L5 cell subtypes were all involved in the upregulation of lipid metabolism, protein and hormone metabolism, and the immune response, respectively. Furthermore, we discovered a novel myofibroblast that expresses COL1A1 and CSN3, has visible epithelial-like characteristics, and shows the potential to differentiate into luminal epithelial cells, especially immune-sensing luminal cells (L5). Conclusions: We constructed the first single-cell atlas of the dairy cow MG, and our new findings of epithelial-like myofibroblast cells and their differentiation trajectories into luminal cells may provide novel insights into the development and lactogenesis in dairy cattle MGs.

scholarly journals Atlas of the human intestine

2020 ◽  
Vol 217 (2) ◽  
Author(s):  
Xi C. He ◽  
Linheng Li
Keyword(s):  
Epithelial Cells ◽  
Single Cell ◽  
Human Colon ◽  
Cell Types ◽  
Nutrient Absorption ◽  
Endocrine Function ◽  
Human Intestine ◽  
Rna Seq

In this issue of JEM, Wang et al. (https://doi.org/10.1084/jem.20191130), using a single-cell RNA-seq approach, establish an atlas of human colon, rectum, and ileum epithelial cells. Their study reveals that different regions have specialized nutrient absorption preferences, microbe defenses, and endocrine function. They also identify new markers for a variety of cell types.

30 A TRANSCRIPTION ACTIVATOR-LIKE EFFECTOR NUCLEASE (TALEN)-MEDIATED UNIVERSAL GENE KNOCK-IN STRATEGY FOR MAMMARY GLANDS-SPECIFIC EXPRESSION OF RECOMBINANT PROTEINS IN DAIRY CATTLE

2014 ◽  
Vol 26 (1) ◽  
pp. 129 ◽  
Author(s):  
S. Lee ◽  
H. Park ◽  
I. Kong ◽  
Z. Wang
Keyword(s):  
Dairy Cattle ◽  
Single Cell ◽  
Recombinant Proteins ◽  
Mammary Glands ◽  
Start Codon ◽  
Regulatory Sequences ◽  
Transcription Activator ◽  
Translation Start Codon ◽  
Casein Gene ◽  
Translation Start

To harness the great capability of producing biologically active recombinant proteins with animal mammary glands, active research has been carried out in the past several decades to develop transgenic animals as bioreactors. However, when a transgene is introduced in the animal genome by random integration, the transgene tends to be subjected to epigenetic silencing, due to the so-called position effect from the chromatin environments surrounding the transgene integration sites, thereby resulting in low-level expression or total suppression. We report a universal transgenic strategy to knock in (KI) transgenes into the bovine β-casein gene locus allowing the expression of a transgene to be totally under the control of the endogenous regulatory sequences of the bovine β-casein gene. This universal KI strategy comprises two key components: one is the design of transcription activator-like effector nuclease (TALEN) constructs targeting the start codon region of bovine β-casein gene, and the other is the design of KI vectors in which a transgene of choice is flanked with homologous arms isolated from the ~500-bp bovine genomic DNAs immediately 5′ and 3′, respectively, of the translation start codon of the bovine β-casein gene. By using the human erythropoietin (hEPO) as the model transgene, we demonstrated that a transgene can be highly efficiently integrated immediately after the translation start codon of the bovine β-casein gene. In brief, the TALEN constructs were assembled by using the Golden Gate protocol. To KI the hEPO transgene, early passage (<5) of fibroblasts established from Holstein dairy cattle were cultured into full confluence in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS), harvested with 0.25% trypsin-EDTA, and co-transfected with KI vector and the TALEN constructs by the Amaxa Nucleofector system. For each experiment, 106 cells were transfected with 5 μg of KI vector and 5 μg of TALEN constructs. After 72 h post-transfection, cells were harvested and subjected to limiting dilution to obtain single-cell derived colonies. To screen for single-cell derived colonies carrying the correctly KI of hEPO in the β-casein locus, we performed genomic PCR amplifying the genomic junctions created by the KI of hEPO gene into the bovine genome. We identified and established 2 hEPO transgenic bovine fibroblast cell lines after screening 10 single-cell derived colonies from the transfected cells (20%). The genotype of these 2 colonies was also confirmed by sequencing the PCR products. We have initiated the effort to produce hEPO transgenic cattle by somatic cell nuclear transfer (SCNT), and the animal cloning results will be reported at the conference.

scholarly journals Single-cell transcriptome analysis reveals differential nutrient absorption functions in human intestine

2019 ◽  
Vol 217 (2) ◽  
Author(s):  
Yalong Wang ◽  
Wanlu Song ◽  
Jilian Wang ◽  
Ting Wang ◽  
Xiaochen Xiong ◽  
...  
Keyword(s):  
Single Cell ◽  
Large Intestine ◽  
Hormone Secretion ◽  
Cell Types ◽  
Nutrient Absorption ◽  
Marker Genes ◽  
Human Intestine ◽  
Colon And Rectum ◽  
Cell Transcriptome ◽  
Single Cell Transcriptome

The intestine plays an important role in nutrient digestion and absorption, microbe defense, and hormone secretion. Although major cell types have been identified in the mouse intestinal epithelium, cell type–specific markers and functional assignments are largely unavailable for human intestine. Here, our single-cell RNA-seq analyses of 14,537 epithelial cells from human ileum, colon, and rectum reveal different nutrient absorption preferences in the small and large intestine, suggest the existence of Paneth-like cells in the large intestine, and identify potential new marker genes for human transient-amplifying cells and goblet cells. We have validated some of these insights by quantitative PCR, immunofluorescence, and functional analyses. Furthermore, we show both common and differential features of the cellular landscapes between the human and mouse ilea. Therefore, our data provide the basis for detailed characterization of human intestine cell constitution and functions, which would be helpful for a better understanding of human intestine disorders, such as inflammatory bowel disease and intestinal tumorigenesis.

Single-cell transcriptomic analyses of dairy cattle ruminal epithelial cells during weaning

Genomics ◽  
2021 ◽  
Author(s):  
Yahui Gao ◽  
Lingzhao Fang ◽  
Ransom L. Baldwin ◽  
Erin E. Connor ◽  
John B. Cole ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Dairy Cattle ◽  
Single Cell

Electron microscopy of keratinocytes cultured on a collagen substrate used as an allograft for the treatment of chronic wounds

1992 ◽  
Vol 50 (2) ◽  
pp. 1104-1105
Author(s):  
Debby A. Jennings ◽  
Michael J. Morykwas ◽  
Louis C. Argenta
Keyword(s):  
Electron Microscopy ◽  
Cell Suspension ◽  
Single Cell ◽  
Chronic Wounds ◽  
Mechanical Stability ◽  
Uv Light ◽  
Type I ◽  
Single Cell Suspension ◽  
Collagen Substrate ◽  
Dermal Collagen

Grafts of cultured allogenic or autogenic keratlnocytes have proven to be an effective treatment of chronic wounds and burns. This study utilized a collagen substrate for keratinocyte and fibroblast attachment. The substrate provided mechanical stability and augmented graft manipulation onto the wound bed. Graft integrity was confirmed by light and transmission electron microscopy.Bovine Type I dermal collagen sheets (100 μm thick) were crosslinked with 254 nm UV light (13.5 Joules/cm2) to improve mechanical properties and reduce degradation. A single cell suspension of third passage neonatal foreskin fibroblasts were plated onto the collagen. Five days later, a single cell suspension of first passage neonatal foreskin keratinocytes were plated on the opposite side of the collagen. The grafts were cultured for one month.The grafts were fixed in phosphate buffered 4% formaldehyde/1% glutaraldehyde for 24 hours. Graft pieces were then washed in 0.13 M phosphate buffer, post-fixed in 1% osmium tetroxide, dehydrated, and embedded in Polybed 812.

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