a7r5 vascular smooth muscle cells  (Millipore)

Journal: bioRxiv

Article Title: Cellular transcriptomics reveals evolutionary adaptation and rumination of vertebrate stomachs

doi: 10.64898/2026.03.09.710455

Figure Lengend Snippet: In situ hybridization of LUC7L transcripts in four stomach chambers (rumen, reticulum, omasum, and abomasum) of sheep and equine stomach, with a scale bar of 100 μm. b, Quantification of the cell proliferation between negative control (NC) and Luc7l -knockdown (si Luc7l ) A7r5 cells, as measured by CCK-8 assay. Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-way ANOVA, where *** indicates P < 0.001. c, Migration of A7r5 cells in a comparison of NC and si Luc7l . Wound healing of A7r5 cells was assessed in images (left) at 36 hours (h) post-scratch, relative to the 0-h time point. Wound edge is highlighted by the dashed white line. Migration rate was calculated and compared between NC and si Luc7l . Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-tailed unpaired t -test, where ** indicates P < 0.01, with a scale bar of 100 μm. d, Cytoskeleton F-actin of A7r5 cells in a comparison of NC (at 0 h) and si Luc7l (at 72 h). The images were all taken from randomly selected fields. Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-tailed unpaired t -test, where ** indicates P < 0.01, with a scale bar of 25 µm. e, Volcano plot showing the up- and down-regulated differentially expressed genes (DEGs) in bulk RNA-seq between the NC and si Luc7l . f, GO cellular component (CC) enrichment analysis of the down-regulated DEGs. g, Schematic diagram illustrating the mechanism that LUC7L regulates the phenotypes of smooth muscle cells (SMCs). The up-regulated expression of LUC7L promotes the switch from synthetic to contractile phenotypes and lowers rates of SMC proliferation and migration. The expression of synthetic genes (e.g., PDGF , FGF , and SERPINE1 genes) are up-regulated significantly in si Luc7l cells, while the contractile genes (e.g., TGF-β and SMAD2 ) are down regulated, compared to NC. h, Representative ventral images of fluorescence emission of abdomen. Mice received a 600 mg/kg body weight of 4-kDa FITC dextran imaging agent in wild-type (WT) mice and homozygous Luc7l knockout ( Luc7l −/− ) mice. Representative ventral images of anesthetized mice are shown at baseline (blank), 5 and 7 min post-gavage. All images were acquired using the same pseudocolor scale of radiance to show relative changes in bioluminescence emission over time. Time to first appearance of fluorescence in the abdomen compared between WT and Luc7l −/− mice. Data are presented as the mean ± SEM (3 biological replicates). P values are determined by two-tailed unpaired t-test, where * indicates P < 0.05. i, Gastric emptying 20 min after gavage of 0.5 ml of phenol red solution in WT and Luc7l −/− mice. Data are presented as the mean ± SEM (6 biological replicates). P values are determined by two-tailed unpaired t -test, where *** indicates P < 0.001.

Article Snippet: A7r5 vascular smooth muscle cells (hereafter A7r5, Procell Life Science & Technology) and STC-1 mouse intestinal enteroendocrine cells (hereafter STC-1, Shanghai Jinyuan Biotechnology Co., Ltd.) were maintained in DMEM (Gibco, 11995073) supplemented with 10% fetal bovine serum (FBS) (Gibco, A5670701) and 1% penicillin-streptomycin (Beyotime, C0222).

Techniques: In Situ Hybridization, Negative Control, Knockdown, CCK-8 Assay, Migration, Comparison, Two Tailed Test, RNA Sequencing, Expressing, Fluorescence, Imaging, Knock-Out

Journal: Nutrients

Article Title: Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

doi: 10.3390/nu16020291

Figure Lengend Snippet: Figure 3. ALP activity in rat aortic A7r5 cells under zinc treatment. Rat aortic A7r5 cells were cultured for 15 d under zinc treatments supplemented with either βGP or NaP as the phosphate source. (A) ALP staining of the ECM using BCIP/NBT as substrate did not show an increase in ALP activity under Zn-deficient conditions with increasing P levels. (B) Measurement of ALP activity using PNPP as substrate did not show a significant effect of zinc (p < 0.05, one-way ANOVA followed by Tukey HSD). (C) Protein expression as assessed through Western blotting showed very weak ALP expression and downregulation under Zn−is only observed in βGP-treated cultures.

Article Snippet: Mouse origin osteoblastic MC3T3-E1 cell subclone 4 (CRL-2593) and rat aortic vascular smooth muscle cells A7r5 (CRL-1444) were obtained commercially from American Type Culture Collection (Manassas, VA, USA).

Techniques: Activity Assay, Cell Culture, Staining, Expressing, Western Blot

Journal: Nutrients

Article Title: Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

doi: 10.3390/nu16020291

Figure Lengend Snippet: Figure 4. Ca and P deposition in zinc-deficient VSMCs supplemented with increasing concentrations of β-GP or NaP. Rat aortic A7r5 cells were cultured for 15 d under zinc treatments supplemented with either βGP or NaP as the phosphate source. (A) Ca deposition was assessed through Alizarin red staining, while (B) P deposition was assessed through von Kossa. Images are representative from n = 4 samples.

Article Snippet: Mouse origin osteoblastic MC3T3-E1 cell subclone 4 (CRL-2593) and rat aortic vascular smooth muscle cells A7r5 (CRL-1444) were obtained commercially from American Type Culture Collection (Manassas, VA, USA).

Techniques: Cell Culture, Staining

Journal: Nutrients

Article Title: Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

doi: 10.3390/nu16020291

Figure Lengend Snippet: Figure 5. Pit1 expression in rat aortic A7r5and osteoblastic MC3T3-E1 cells under zinc treatment. Both A7r5 and MC3T3-E1 cells were cultured up to 15 d under zinc treatments. GM (normal growth media) was used as control in A7r5 while OSM (normal osteogenic medium) was used as control for MC3T3-E1 cells. Data represent means ± SD (n = 3). The statistical significance between values for each group was assessed through Dunnett’s t-test. * p < 0.05 between the Zn−or Zn + and control groups.

Article Snippet: Mouse origin osteoblastic MC3T3-E1 cell subclone 4 (CRL-2593) and rat aortic vascular smooth muscle cells A7r5 (CRL-1444) were obtained commercially from American Type Culture Collection (Manassas, VA, USA).

Techniques: Expressing, Cell Culture, Control

Journal: Nutrients

Article Title: Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

doi: 10.3390/nu16020291

Figure Lengend Snippet: Figure 6. Mineral measurement in extracellular (media) and intracellular (cell) fractions. Aortic A7r5 cells were cultured up to 15 d under zinc treatments. (A) The concentrations of P in both media and cell fractions were not significantly affected by zinc. Upon addition of PFA, media P increased under Zn−while cellular P decreased in Zn−and Zn+. (B) Ca concentrations were also not affected by zinc and, like P, its concentration decreased upon PFA addition. (C) The levels of zinc in the media confirmed our zinc treatment, although the cellular zinc levels remained unchanged. The addition of PFA did not alter cellular zinc levels. The mean with the different superscripts are significantly different by Zn treatment. p < 0.05, one-way ANOVA followed by post hoc Tukey HSD test.

Article Snippet: Mouse origin osteoblastic MC3T3-E1 cell subclone 4 (CRL-2593) and rat aortic vascular smooth muscle cells A7r5 (CRL-1444) were obtained commercially from American Type Culture Collection (Manassas, VA, USA).

Techniques: Cell Culture, Concentration Assay

Journal: Nutrients

Article Title: Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

doi: 10.3390/nu16020291

Figure Lengend Snippet: Figure 7. Inhibition of phosphate uptake via Pit1. Both A7r5 and MC3T3-E1 cells were cultured up to 15 d under zinc treatments. (A) Inhibition of phosphate uptake did not inhibit Pit1 expression in both A7r5 VSMCs and osteobalstic MC3T3-E1 cells. Data represent means ± SD (n = 3). The statistical significance between values for each group was assessed through Dunnett’s t-test. * p < 0.05 between Zn−or Zn + and control groups. (B) However, Ca and P deposition in both cells lines were significantly inhibited by treatment with PFA. (C). In A7r5 VSMCs, treatment with PFA restored the expression of the VSMC markers SM22α and calponin under zinc deficiency.

Article Snippet: Mouse origin osteoblastic MC3T3-E1 cell subclone 4 (CRL-2593) and rat aortic vascular smooth muscle cells A7r5 (CRL-1444) were obtained commercially from American Type Culture Collection (Manassas, VA, USA).

Techniques: Inhibition, Cell Culture, Expressing, Control

Journal: Nutrients

Article Title: Zinc Deficiency Promotes Calcification in Vascular Smooth Muscle Cells Independent of Alkaline Phosphatase Action and Partly Impacted by Pit1 Upregulation.

doi: 10.3390/nu16020291

Figure Lengend Snippet: Figure 8. Schematic summary of the molecular mechanisms enabled through Pit1 expression in VSMCs; this is a target for zinc deficiency and may mediate the inhibition of VSMC marker expression under zinc deficiency.

Article Snippet: Mouse origin osteoblastic MC3T3-E1 cell subclone 4 (CRL-2593) and rat aortic vascular smooth muscle cells A7r5 (CRL-1444) were obtained commercially from American Type Culture Collection (Manassas, VA, USA).

Techniques: Expressing, Inhibition, Marker