Journal: The Journal of clinical endocrinology and metabolism

Article Title: Coexpression of fractalkine and its receptor in normal human endometrium and in endometrium from users of progestin-only contraception supports a role for fractalkine in leukocyte recruitment and endometrial remodeling.

doi: 10.1210/jc.2003-031379

Figure Lengend Snippet: FIG. 1. Immunohistochemical localization of fractalkine (A–H and R–T) in human endometrium. A, Proliferative phase endometrium showing diffuse immunostaining for fractalkine in glandular epithelium (GE) and leukocytes (). More intense staining was seen in the early secretory (B), midsecretory (C), and late secretory (D and E) phases. Staining appeared to be in vesicles (V) in the basal (ba) region in GE in the early secretory phase (B), more diffuse in the midsecretory phase (C), and apically located (ap) in the late secretory phase (D and inset), when strong staining was also observed in blood vessels (bv) and decidualizing stroma (Dec) surrounding the spiral arterioles (E). Fractalkine was also identified in decidua (Dec) in early pregnancy (F). No staining was observed after preadsorption of antibody with fractalkine peptide (E and inset). Fractalkine-staining leukocytes were identified by colocalization studies on adjacent 2-m sections. Staining pairs are fractalkine (G)

Article Snippet: Negative controls were also included, where the primary antibody was preabsorbed with 1 g/ml fractalkine blocking peptide (SC-7226, Santa Cruz Biotechnology) at 4 C for 48 h. A section from a single block of endometrial tissue was included in each staining run for quality control.

Techniques: Immunohistochemical staining, Immunostaining, Staining

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Coexpression of fractalkine and its receptor in normal human endometrium and in endometrium from users of progestin-only contraception supports a role for fractalkine in leukocyte recruitment and endometrial remodeling.

doi: 10.1210/jc.2003-031379

Figure Lengend Snippet: FIG. 2. Fractalkine (FKN) immunostaining intensity in human en- dometrium, across the menstrual cycle and in early pregnancy. A, Semiquantitative scoring of FKN in glandular epithelium and stro- mal cells in the following tissues: menstrual (ME; n 5), proliferative (PR; n 7), early secretory (ES; n 6), midsecretory (MS; n 6), late secretory (LS; n 6), and early pregnancy (PREG; n 10). Results shown are the mean SEM. *, P 0.05 (in ES, LS, and PREG, glandular immunostaining levels are significantly elevated compared PR and MS levels). ††, P 0.01 (in LS, stromal immunostaining levels are significantly elevated compared with MS levels). B, Semiquan- titative scoring of FKN in leukocytes. Results shown are the mean SEM.

Article Snippet: Negative controls were also included, where the primary antibody was preabsorbed with 1 g/ml fractalkine blocking peptide (SC-7226, Santa Cruz Biotechnology) at 4 C for 48 h. A section from a single block of endometrial tissue was included in each staining run for quality control.

Techniques: Immunostaining

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Coexpression of fractalkine and its receptor in normal human endometrium and in endometrium from users of progestin-only contraception supports a role for fractalkine in leukocyte recruitment and endometrial remodeling.

doi: 10.1210/jc.2003-031379

Figure Lengend Snippet: FIG. 4. Semiquantitative scoring of glandular epithelium, stromal, and leukocyte immunostaining levels for fractalkine (FKN) and CX3CR1 in human endometrium exposed to P-only contraceptives. A, FKN localization in endometrium before insertion of LNG-IUS in the proliferative phase (PR; n 4) or secretory phase (SEC; n 5) and at 3, 6, and 12 months postinsertion (n 3/group). *, P 0.05, significant increase in FKN-immunopositive leukocytes 3 months postinsertion compared with preinsertion controls. Epithelial immu- nostaining was absent 3 months postinsertion and significantly de- creased 12 months postinsertion (*, P 0.05). Stromal cells were strongly positive for FKN at all stages postinsertion (*, P 0.05). B, FKN immunostaining in endometrium from users of Depo-Provera (DEPO) and Norplant (NORP) compared with nonusers (controls) from proliferative phase (PR; n 7) and secretory phase (SEC; n

Article Snippet: Negative controls were also included, where the primary antibody was preabsorbed with 1 g/ml fractalkine blocking peptide (SC-7226, Santa Cruz Biotechnology) at 4 C for 48 h. A section from a single block of endometrial tissue was included in each staining run for quality control.

Techniques: Immunostaining

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Coexpression of fractalkine and its receptor in normal human endometrium and in endometrium from users of progestin-only contraception supports a role for fractalkine in leukocyte recruitment and endometrial remodeling.

doi: 10.1210/jc.2003-031379

Figure Lengend Snippet: FIG. 5. Diagrammatic illustration of fractalkine expression and its potential functional significance. Epithelial (E) and stromal (S) fractalkine (FKN) levels are maximal during the secretory phase of the menstrual cycle. In the midsecretory (MS) phase, there is a dramatic influx of macrophages; FKN produced by E and S could be involved in their positioning and activation. This also coincides with the time of blastocyst implantation; FKN could be acting to increase blastocyst/epithelium receptivity. Production of FKN by endometrial vasculature only occurs premenstrually. Macrophages and neutrophils express CX3CR1; therefore, endometrial FKN could be involved in their selective recruitment, positioning, and activation for the initiation of menstruation. Prolif, Proliferative phase; LS, late secretory phase; V, fractalkine positive vessel staining.

Article Snippet: Negative controls were also included, where the primary antibody was preabsorbed with 1 g/ml fractalkine blocking peptide (SC-7226, Santa Cruz Biotechnology) at 4 C for 48 h. A section from a single block of endometrial tissue was included in each staining run for quality control.

Techniques: Expressing, Functional Assay, Produced, Activation Assay, Staining

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Cytoplasmic expression of fibroblast growth factor receptor-4 in human pituitary adenomas: relation to tumor type, size, proliferation, and invasiveness.

doi: 10.1210/jc.2003-031489

Figure Lengend Snippet: FIG. 1. Detection of FGFR4 reactivity in pituitary cells. A–D, Immunohistochem- ical localization of FGFR4 in human pi- tuitary adenomas. C-terminal FGFR4 immunoreactivity is seen in the cyto- plasm of tumor cells in varied tumor types. A, GH cell adenoma. B, ACTH cell adenoma. C, FSH/LH adenoma. D, Null cell adenoma. The immunoreactivity of FGFR4 is variable but always very in- tense throughout the cytoplasm (original magnification 50–100). E and F, Im- munohistochemical localization of FGFR4 and ptd-FGFR4 in transfected pituitary cells. Rat pituitary tumor-derived GH4 cells were stably transfected with full- length FGFR4 (E) or ptd-FGFR4 (F). Im- munocytochemical examination was per- formed using an antibody that recognizes the C terminus of FGFR4. Note the pre- dominant cytoplasmic pattern of staining in ptd-FGFR4 transfected cells, com- pared with the membrane reactivity in wild-type FGFR4-transfected cells. G, Western blot detection of FGFR4 in hu- man pituitary adenomas. Protein lysates from seven human gonadotroph adeno- mas were electrophoresed and blotted with an antibody that recognizes the C terminus of FGFR4. The extreme right lane contains a positive control from HEK293 cells transiently transfected with ptd-FGFR4 (upper panel). Preab- sorption of the primary antibody with pu- rified antigen abolishes the lower 65-kDa protein (lower panel). The band migrat- ing just above 65 kDa is a nonspecific species that is not abolished by preab- sorption.

Article Snippet: Furthermore, the specificity of all reactions for FGFR4 was verified by replacing the primary antibody with normal serum, examining negative control tissues, and preabsorbing primary antibody with purified FGFR4 peptide (Santa Cruz, antibody: peptide/6:1).

Techniques: Transfection, Derivative Assay, Stable Transfection, Staining, Membrane, Western Blot, Positive Control

Journal: The Journal of clinical endocrinology and metabolism

Article Title: Cytoplasmic expression of fibroblast growth factor receptor-4 in human pituitary adenomas: relation to tumor type, size, proliferation, and invasiveness.

doi: 10.1210/jc.2003-031489

Figure Lengend Snippet: FIG. 2. Cytoplasmic expression levels of FGFR4 and Ki-67 LI in pituitary ad- enomas. A, The expression levels of FGFR4 in macroadenomas are signifi- cantly higher than those in microadeno- mas (P 0.02). B, The expression level of FGFR4 in invasive adenomas is not significantly different from noninvasive adenomas. C, The mean Ki-67 LI (per- cent) was significantly higher in tumors that express high levels of FGFR4 than in the low FGFR4-expressing group, which in turn was higher than in the FGFR4-negative group of tumors (P 0.03; P 0.002, respectively). D and E, Ki-67 LI was higher in macroadenomas and invasive adenomas than in mi- croadenomas and noninvasive adeno- mas (P 0.05; P 0.01, respectively).

Article Snippet: Furthermore, the specificity of all reactions for FGFR4 was verified by replacing the primary antibody with normal serum, examining negative control tissues, and preabsorbing primary antibody with purified FGFR4 peptide (Santa Cruz, antibody: peptide/6:1).

Techniques: Expressing

Journal: Oncology reports

Article Title: Expression of angiopoietin-like 4 (ANGPTL4) in human colorectal cancer: ANGPTL4 promotes venous invasion and distant metastasis.

doi: 10.3892/or.2011.1176

Figure Lengend Snippet: Figure 1. Immunohistochemical staining for ANGPTL4 in human tissues. (A) Faintly expression in the surface lining cells of normal colon mucosa (arrow- heads), (B) strongly in cytoplasm of moderately-differentiated adenocarcinoma, and (C) poorly-differentiated adenocarcinoma (non-solid type) of human colorectal cancer (magnification, x100).

Article Snippet: Deparaffinized sections were preincubated with normal rabbit serum to prevent non-specific binding, and then incubated overnight at 4 ̊C with an optimal dilution (0.1 μg/ml) of a primary polyclonal goat antibody against human Angptl4 (r&D systems, Inc., minneapolis, mA, usA). the slides were sequentially incubated with a biotinylated rabbit antigoat immunoglobulin antibody, and the reaction products were viewed using diaminobenzidine (DAb; Dako ltd.) and counterstained with hematoxylin. primary antibody preabsorbed with excess recombinant Angptl4 peptides (r&D systems, Inc.) was used as negative control.

Techniques: Immunohistochemical staining, Staining, Expressing

Journal: Oncology reports

Article Title: Expression of angiopoietin-like 4 (ANGPTL4) in human colorectal cancer: ANGPTL4 promotes venous invasion and distant metastasis.

doi: 10.3892/or.2011.1176

Figure Lengend Snippet: Figure 2. Overall survival based on the expression of ANGPTL4 in human colorectal cancer (60 cases). ANGPTL4 expression was not associated with the overall survival in univariate survival analysis. (p=0.455 in log-rank test).

Article Snippet: Deparaffinized sections were preincubated with normal rabbit serum to prevent non-specific binding, and then incubated overnight at 4 ̊C with an optimal dilution (0.1 μg/ml) of a primary polyclonal goat antibody against human Angptl4 (r&D systems, Inc., minneapolis, mA, usA). the slides were sequentially incubated with a biotinylated rabbit antigoat immunoglobulin antibody, and the reaction products were viewed using diaminobenzidine (DAb; Dako ltd.) and counterstained with hematoxylin. primary antibody preabsorbed with excess recombinant Angptl4 peptides (r&D systems, Inc.) was used as negative control.

Techniques: Expressing

Journal: Oncology reports

Article Title: Expression of angiopoietin-like 4 (ANGPTL4) in human colorectal cancer: ANGPTL4 promotes venous invasion and distant metastasis.

doi: 10.3892/or.2011.1176

Figure Lengend Snippet: Figure 3. RT-PCR and Western blot analysis for ANGPTL4 in human colo rectal cancer: tissues and cultured cell lines. (A) RT-PCR. M, 100 bp ladder marker (Invitrogen, Inc.). (B) Western blot analysis. Human colorectal cancer tissues (1-6) and human colorectal cancer cell lines (7-12). [1, Cancer tissue; cases 1 and 2, normal gastric tissue; cases 1 and 3, cancer tissue; cases 2 and 4, normal gastric tissue; cases 2, 5, cancer tissue; cases 3 and 6, normal gastric tissue; cases-3 and 7, Caco-2; 8, Colo201; 9, Colo320DM; 10, DLD-1; 11, WiDr,; 12, LS123].

Article Snippet: Deparaffinized sections were preincubated with normal rabbit serum to prevent non-specific binding, and then incubated overnight at 4 ̊C with an optimal dilution (0.1 μg/ml) of a primary polyclonal goat antibody against human Angptl4 (r&D systems, Inc., minneapolis, mA, usA). the slides were sequentially incubated with a biotinylated rabbit antigoat immunoglobulin antibody, and the reaction products were viewed using diaminobenzidine (DAb; Dako ltd.) and counterstained with hematoxylin. primary antibody preabsorbed with excess recombinant Angptl4 peptides (r&D systems, Inc.) was used as negative control.

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Cell Culture, Marker

Journal: Biology of reproduction

Article Title: Inducible nitric oxide synthase in the rat testis: evidence for potential roles in both normal function and inflammation-mediated infertility.

doi: 10.1095/biolreprod63.5.1285

Figure Lengend Snippet: FIG. 1. A representative immunohistochemical distribution of iNOS protein in the normal and LPS-treated rat testis. A) Negative control section from a high dose LPS-treated rat testis 6 h after LPS injection probed with the iNOS antiserum preabsorbed with the immunizing peptide. B–E) A normal rat testis probed with the iNOS antiserum. Within the normal rat testis iNOS protein was found in low levels within discrete populations of elongating spermatids and was most easily visible in step 9 spermatids (et, B), and pachytene spermatocytes (pc, C). Inducible NOS immunostaining was also seen within peritubular cells at most stages of the seminiferous cycle (pt, C) with the exception of stage XII tubules (D) and in low levels within Sertoli cell cytoplasm (sc, D). The majority of staining within the normal testis, however, was confined to the Leydig cells (lc, D and E). Testicular macrophages (m) were not stained for iNOS protein and were distinguished from Leydig cells using double labeling for iNOS and the resident macrophage marker ED2 (E) (b, blood vessel). F–H) Following injection with LPS an increase in iNOS immunostaining was seen within germ cells (F), Sertoli cells (G), and Leydig cells (H). Rats were killed 6 h following injection with 5 mg/kg LPS. The cell marked by an arrowhead in H is a resident testicular macrophage that did not express iNOS protein in response to LPS injection. Magnifications: A, B) 340, B, C, G, H) 31000, D) 3200, and E) 3100. Bars 5 25 mm.

Article Snippet: The specificity of the immunohistochemical staining in our hands was established using either nonimmune rabbit sera in place of the primary antibody solution or by preabsorbing the iNOS antiserum using the iNOS immunogen peptide, as outlined by the manufacturer (Santa Cruz).

Techniques: Immunohistochemical staining, Negative Control, Injection, Immunostaining, Staining, Labeling, Marker