Fig 1: Immunohistochemical staining for SPOCK1 in various subtypes of breast carcinoma.SPOCK1 was not expressed in representative cases of invasive lobular carcinoma (A), mucinous carcinoma (B), invasive papillary carcinoma, (C) and invasive micropapillary carcinoma (D). SPOCK1 expression in vascular smooth muscle wall (arrow) served as internal positive control. Two representative SPOCK1-positive cases of metaplastic carcinoma are shown (E and F). Intense SPOCK1 expression was observed in both carcinomatous (Ca) and sarcomatous (Sa) elements in a representative biphasic metaplastic carcinoma (E). SPOCK1 was strongly stained in one representative monophasic epithelial metaplastic carcinoma (squamous metaplasia) (F). (Magnification ×200).
Fig 2: SPOCK1 was a TGF-ß-induced myoepithelial marker and SPOCK1 overexpression enhanced invasiveness.(A) Immunohistochemistry demonstrated that SPOCK1 was the only myoepithelial marker among the evaluated TGF-ß–upregulated gene products in MCF10A cells. CXCR7 staining was observed in luminal epithelia but not myoepithelia whereas FBN1 staining was observed diffusely in the stroma but not in the epithelia. IGFL2, PRRX2, PLAT, PCDH9, POSTN and NOV staining was not found in the ductolobular units (left and middle panels). By contrast, SPOCK1 was immunolocalized within (arrow) or beneath (arrowhead) the myoepithelia (right panel). (Magnification × 400). (B) Upregulation of SPOCK1 at mRNA levels (upper panels) and protein levels (lower panels) were observed in MCF10A, M10, and MCF12A cells, 3 days after treatment with TGF-ß. S26 was used as an mRNA loading control and GAPDH was used as a protein loading control. (C) Western blotting was used to detect the expression of SPOCK1 in MCF10A, T47D, and MB231 cells 24 h after transient transfection with pCDH-SPOCK1 or control vector pCDH. (D) The invasive capability and proliferation were measured in the cells shown in (C). Data from invasion assay are shown as the mean ± SD of 3 fields. Data from MTT assay are shown as mean ± SD of 3 independent experiments. These results are presented as the percentage relative to their control cells (*, P < 0.05; N.S., nonsignificant).
Fig 3: SPOCK1 expression and Kaplan–Meier analysis of overall survival in IDC patients.(A) Representative examples of IDC which were positive (left) and negative (right) for SPOCK1 expression. Note that the SPOCK1 expression in vascular smooth muscle wall (arrow) serve as internal positive control (magnification ×400). (B) SPOCK1 expression in IDCs was significantly associated with decreased overall survival (P = 0.001, log-rank test).
Fig 4: A working model shows the molecular mechanism underlying the ability of SPOCK1 to promote the progression of clear cell renal cell carcinoma (ccRCC) cells. The prometastatic effect of SPOCK1 on ccRCC cells was attributed to induce the upregulation of Snail family members (Snail and Slug) following transcriptional downregulation of E-cadherin and upregulation of the matrix metalloproteinase (MMP)-2. Moreover, SPOCK1-induced upregulation of Snail family members also shows the positive feedback regulation of SPOCK1. These signal pathways ultimately trigger epithelial-to-mesenchymal transition (EMT) progression and subsequent promotion of ccRCC metastasis. Green ovals indicate hypothetical regulators which might be involved in the SPOCK1-mediated interplay of Akt and Snail family members.
Fig 5: Clinical significance of SPOCK1 in clear cell renal cell carcinoma (RCC; ccRCC). (A) SPOCK1 gene expression levels in RCC specimens (n = 69), ccRCC specimens (n = 32), and normal tissue samples (n = 23) were measured by Affymetrix oligonucleotide arrays obtained from the GEO (GSE15641). (B) SPOCK1 gene expression levels in ccRCC samples from TCGA were compared according to the clinical stage, tumor size (T stage), lymph node metastasis (N stage), and distal metastasis (M stage). Statistical significance was analyzed by a t-test. * p < 0.05, *** p < 0.001. (C) Kaplan–Meier curves for survival of patients with ccRCC or papillary (p)RCC, as categorized according to high or low expression of SPOCK1. The p value indicates a comparison between patients with SPOCK1high and SPOCK1low. The ccRCC and pRCC datasets were retrieved from TCGA. (D) SPOCK1 protein expression levels in ccRCC specimens and adjacent normal tissue samples or benign tumor (oxyphilic adenoma) samples were measured by IHC staining. The right panels are the enlarged images of left panels. Scale bars of left panel and right panel are 200 and 100 µM, respectively.
Supplier Page from MilliporeSigma for Anti-SPOCK1 antibody produced in rabbit