anti collageniv  (SouthernBiotech)

Journal: BMC Developmental Biology

Article Title: Smad4-dependent pathways control basement membrane deposition and endodermal cell migration at early stages of mouse development

doi: 10.1186/1471-213X-9-54

Figure Lengend Snippet: Mature Smad4 mutant embryoid bodies display a thickened endoderm and expanded basement membrane . The day 10 Smad4 null EBs strongly express Dab2. The thick extracellular matrix, detectable by hematoxylin and eosin (H&E) staining, stains positive for both laminin and collagenIV.

Article Snippet: Slides were then incubated with primary antibodies as follows, anti-laminin (Sigma, L-9393; 1:200); anti-collagenIV (Chemicon, AB756P; 1:100) anti-Oct4 (Santa Cruz sc-8628; 1:200).

Techniques: Mutagenesis, Staining

Journal: bioRxiv

Article Title: GM-CSF drives immune-mediated glomerular disease by licensing monocyte-derived cells to produce MMP12

doi: 10.1101/2022.06.13.495915

Figure Lengend Snippet: (A) Unsupervised clustering and UMAP of FACS-analysis of renal leukocytes from nephritic wild-type and GM-CSF/ Cf2 −/− mice. Cell counts in the respective clusters from wildtype and GM-CSF/ Csf2 -/- animals. (B) Unsupervised clustering and UMAP dimensional reduction of scRNA-seq of renal leukocytes from wildtype and GM-CSF/ Csf2 −/− mice ten days after cGN induction. Clusters were annotated according to their gene expression profiles. Fraction of cells in the respective clusters from wildtype and GM-CSF/ Cf2 −/− animals. (C) Heatmap of key marker gene expression of the indicated clusters form scRNA-seq. (D) RT-PCR analysis of renal Mmp12 expression in wildtype and GM-CSF −/− mice. (E) Combined immunofluorescence staining of CD3 + (green), MAC2 + (turquoise), and FISH ( Mmp12 , red) of renal leukocytes in wildtype and GM-CSF/ Csf2 −/− mice. (F) Pathway analysis of differentially expressed genes of renal MdCs from nephritic wildtype and GM-CSF/ Csf2 −/− mice (color code indicating the log2-fold change in wildtype versus GM-CSF/ Csf2 −/− animals). (G) FACS sorted MdC like cells from the spleen of control mice were in vitro cultured for 24 hours with/without addition of recombinant mouse GM-CSF. Mmp12 gene expression and protein expression in the supernatant were quantified. (H) FACS sorted MdCs from kidneys of cGN mice were in vitro cultured for 24 hours with/without addition of recombinant mouse GM-CSF. Mmp12 gene expression and protein expression in the supernatant were quantified. (I) FACS sorted MdC like cells from spleens of control mice were in vitro cultured for 1 hour with/without addition of recombinant mouse GM-CSF and with addition of indicated pharmacological inhibitors. Mmp12 gene expression was quantified by RT-PCR. (J-M) Crescentic GN induction and treatment with pharmacological MMP12 inhibitor or vehicle. (J) cGN was induced in wildtype mice and MMP12 inhibitor was given by oral gavage at indicated time points (bis in die, B.I.D.). (K) Quantification of glomerular crescents, tubulointerstitial damage, and BUN levels. (L) Representative confocal micrographs depicting extent of glomerular injury in kidneys of mice with/without MMP12 inhibitor treatment. Note the strong fibrin/fibrinogen signal within the capillaries of the cGN + vehicle glomerulus, whereas msIgG and shIgG/CollagenIV signals at the glomerular filtration barrier are comparable between both groups. (M) Quantification of fibrin positive glomeruli detected by immunohistochemistry. (N) cGN was induced in wildtype mice and GM-CSF-deficient mice as well as MMP12 inhibitor was given by oral gavage at indicated time points (bis in die, B.I.D.). (O) Quantification of glomerular crescents, tubulointerstitial damage, and BUN levels. Symbols represent individual data points with the mean as a bar (* P<0.05, ** P<0.01, *** P<0.005, **** p<0.001). Data are representative of two independent experiments.

Article Snippet: For immunofluorescence, the following primary antibodies were incubated overnight in blocking buffer at 4°C: CD68 (KP1, Invitrogen, Invitrogen, Carsbad, CA), CD3 (A0452, Dako, Glostrup, Denmark), CXCR3 (1C6, BD Biosciences), Mac-2 (M3/38, Cedarlane, Burlington, Canada), CXCR3 (Bioss Antibodies, Woburn, MA), MHCII (sc-59322, Santa Cruz), Fibrin/Fibrinogen (A0080, Dako), CollagenIV (1340-01, SouthernBiotech).

Techniques: Gene Expression, Marker, Reverse Transcription Polymerase Chain Reaction, Expressing, Immunofluorescence, Staining, Control, In Vitro, Cell Culture, Recombinant, Filtration, Immunohistochemistry

Journal: IBRO Neuroscience Reports

Article Title: Early mobilization in spinal cord injury promotes changes in microglial dynamics and recovery of motor function

doi: 10.1016/j.ibneur.2022.04.002

Figure Lengend Snippet: A, B: Merged Images of quadruple staining with GFAP (green), GAP43 (white), and collagenⅣ (red) at the spinal cord injury lesion. Image of the 3-week EM group spinal cord lesion site overview is shown in A. Scale bar: 200 µm. The lesion center is enclosed by the white dotted line. B: Enlargement image of spinal cord lesion. Yellow arrows indicate representative GAP43 structures (white). The border of lesion center is indicated by the yellow dotted line. Scale bars: 50 µm. C : Comparison of the area of GAP43 structures and lesion center between the 3-week NEM and 3-week EM groups. Data are presented as mean ± SEM. * p-value= 0.011. D : Comparison of the GAP43 density of lesion center between the 3-week NEM and 3-week EM groups. Data are presented as mean ± SEM. * p-value= 0.016.

Article Snippet: Mouse monoclonal antibodies against GAP43 (1:500; ab12990, Abcam), rabbit polyclonal antibodies against collagenIV (1:100; ab6586, Abcam), and goat polyclonal antibodies against glial fibrillary acidic protein (GFAP; 1:100; ab53554, Abcam) were used as primary antibodies to label growth cones, collagen fibers, and astrocytes, respectively.

Techniques: Staining, Comparison