Crucial Role for Endothelial Cell α2ß1 Integrin Receptor Clustering in Collagen-Induced Angiogenesis.

Angiogenesis is a crucial mechanism of vascular growth and regeneration that requires biosynthesis and cross-linking of collagens in vivo and is induced by collagen in vitro. Here, we use an in vitro model in which apical Type I collagen gels rapidly induce angiogenesis in endothelial monolayers. We extend previous studies demonstrating the importance of the endothelial α2ß1 integrin, a key collagen receptor, in angiogenesis by investigating the roles of receptor clustering and conformational activation. Immunocytochemical localization of α2ß1 integrins in endothelial monolayers showed a concentration of integrins along cell-cell borders. After inducing angiogenesis with collagen, the receptors redistributed to apical cell surfaces, aligning with collagen fibers, which were also redistributed during angiogenesis. Levels of conformationally activated α2ß1 integrins were unchanged during angiogenesis and undetected on endothelial cells binding collagen in suspension. We mimicked the polyvalency of collagen fibrils using antibody-coated polystyrene beads to cluster endothelial cell surface α2ß1 integrins, which induced rapid angiogenesis in the absence of collagen gels. Clustering of αvß3 integrins and PECAM-1 but not of α1 integrins also induced angiogenesis. Soluble antibodies alone had no effect. Thus, the angiogenic property of collagen may reside in its ability to ligate and cluster cell surface receptors such as α2ß1 integrins. Furthermore, synthetic substrates that promote the clustering of select endothelial cell surface receptors mimic the angiogenic properties of Type I collagen and may have applications in promoting vascularization of engineered tissues. Anat Rec, 2019. © 2019 American Association for Anatomy.

MRI of prostatic urethral mucinous urothelial carcinoma: Expanding the differential diagnosis for T2 hyperintense prostatic masses.

We report the case of a 66-year-old previously healthy man presenting with blood and mucus in his urine. Cystoscopy revealed a mass in the prostatic urethra, and endoscopic biopsy showed adenocarcinoma in situ with mucinous features. Endorectal multiparametric prostate MRI demonstrated a 1.9 cm T2 hyperintense mass in the peripheral zone of the left prostatic apex with extension into the urethral lumen. No diffusion restriction or early enhancement was seen in the mass. Radical prostatectomy was performed, and final pathology demonstrated a mucin-producing urothelial adenocarcinoma arising from the prostatic urethra. The peripheral zone T2 hyperintense abnormality correlated with abundant pools of mucin extending into the prostatic stroma and surrounded by neoplastic prostatic glandular cells. We conclude prostatic urethral mucinous urothelial carcinoma should be included in the differential diagnosis for T2 hyperintense prostatic masses.

Effects of Estrogen on Spermatogenesis in Transgender Women.

OBJECTIVE: To characterize spermatogenesis in the estrogenized transgender patient. MATERIALS AND METHODS: This is a retrospective, single-center, cross-sectional study. Seventy-two transgender women underwent gender-affirming orchiectomy between May 2015 and January 2017. All were on long-term (>1 year) cross-sex hormonal therapy prior to orchiectomy. Patient data were obtained via chart review. Histologic analysis was performed by a pathology resident under the supervision of a genitourinary pathologist. The main outcome is histologic presence of germ cells and presence of spermatids (a proxy for preserved spermatogenesis) in orchiectomy specimens. RESULTS: There were 141 pathologic specimens available for analysis. Germ cells were present in 114 out of 141 (81%) testicles. Spermatids were present in 57 (40%) testicles. Presence of germ cells was associated with older age (43 vs 35 years, P = .007) and increased testicular weight (28.6 g vs 19.3 g, P <.001). Presence of spermatids was associated with increased weight (31.5 g vs 23.3 g, P <.001) and volume (20.3 mL vs 12.6 mL, P <.001). There was a linear correlation between testis volume and preserved spermatogenesis (Pearson's r = 0.448, P <.001). CONCLUSION: Despite long-term hormone therapy, the majority (80%) of transgender women have germ cells present in the testicle. Spermatogenesis is preserved in approximately 40% of these individuals. Duration of hormonal therapy did not affect the degree of preservation of germ cells or spermatogenesis but starting hormonal treatment at a younger age may be associated with decreased germ cells in the testicle. Volume of testicles predict presence of preserved spermatogenesis.

Non-enzymatic glycation of type I collagen diminishes collagen-proteoglycan binding and weakens cell adhesion.

Non-enzymatic glycation of type I collagen occurs in aging and diabetes, and may affect collagen solubility, charge, polymerization, and intermolecular interactions. Proteoglycans(1) (PGs) bind type I collagen and are proposed to regulate fibril assembly, function, and cell-collagen interactions. Moreover, on the collagen fibril a keratan sulfate (KS) PG binding region overlaps with preferred collagen glycation sites. Thus, we examined the effect of collagen modified by simple glycation on PG-collagen interactions. By affinity coelectrophoresis (ACE), we found reduced affinities of heparin and KSPGs for glycated but not normal collagen, whereas the dermatan sulfate (DS)PGs decorin and biglycan bound similarly to both, and that the affinity of heparin for normal collagen decreased with increasing pH. Circular dichroism (CD) spectroscopy revealed normal and glycated collagens to assume triple helical conformations, but heparin addition caused precipitation and decreased triple helical content-effects that were more marked with glycated collagen. A spectrophotometric assay revealed slower polymerization of glycated collagen. However, ultrastructural analyses indicated that fibrils assembled from normal and glycated collagen exhibited normal periodicity, and had similar structures and comparable diameter distributions. B-cells expressing the cell surface heparan sulfate PG syndecan-1 adhered well to normal but not glycated collagen, and endothelial cell migration was delayed on glycated collagen. We speculate that glycation diminishes the electrostatic interactions between type I collagen and PGs, and may interfere with core protein-collagen associations for KSPGs but not DSPGs. Therefore in vivo, collagen glycation may weaken PG-collagen interactions, thereby disrupting matrix integrity and cell-collagen interactions, adhesion, and migration.

Dominant intraprostatic cancer confirmed by direct MRI-guided biopsy: Concordance with histopathological findings.

PURPOSE: To investigate the concordance between dominant intraprostatic cancer seen on endorectal multiparametric MRI and confirmed by MRI-targeted biopsy with histopathological findings at radical prostatectomy, since existing literature has emphasized the miss rather than the concordance rate of MRI. MATERIALS AND METHODS: We retrospectively identified 20 patients who underwent radical prostatectomy after a dominant intraprostatic cancer focus was identified at endorectal multiparametric MRI and confirmed by MRI-targeted biopsy. Concordance was determined by comparing the location and Gleason grade group of dominant tumor at MRI with the location and Gleason grade group determined at histopathological review. RESULTS: Mean patient age was 65 years (range, 48 to 76) and median serum prostatic specific antigen level was 9.4 ng/mL (range, 4.6 to 58.0). In all 20 patients, the location of dominant tumor based on MRI and targeted biopsy corresponded with the dominant tumor location at histopathology. In 9 patients, Gleason grade group was the same at targeted biopsy and final histopathology. In 9 patients, final Gleason grade group was higher and in two patients it was lower. CONCLUSION: Our preliminary results suggest dominant tumor as determined by endorectal multiparametric MRI and confirmed by a positive MRI-targeted biopsy has high concordance with histopathological findings at radical prostatectomy for location, and reasonable concordance for Gleason grade group.

SPOP mutations in prostate cancer across demographically diverse patient cohorts.

BACKGROUND: Recurrent mutations in the Speckle-Type POZ Protein (SPOP) gene occur in up to 15% of prostate cancers. However, the frequency and features of cancers with these mutations across different populations is unknown. OBJECTIVE: To investigate SPOP mutations across diverse cohorts and validate a series of assays employing high-resolution melting (HRM) analysis and Sanger sequencing for mutational analysis of formalin-fixed paraffin-embedded material. DESIGN SETTING AND PARTICIPANTS: 720 prostate cancer samples from six international cohorts spanning Caucasian, African American, and Asian patients, including both prostate-specific antigen-screened and unscreened populations, were screened for their SPOP mutation status. Status of SPOP was correlated to molecular features (ERG rearrangement, PTEN deletion, and CHD1 deletion) as well as clinical and pathologic features. RESULTS AND LIMITATIONS: Overall frequency of SPOP mutations was 8.1% (4.6% to 14.4%), SPOP mutation was inversely associated with ERG rearrangement (P<.01), and SPOP mutant (SPOPmut) cancers had higher rates of CHD1 deletions (P<.01). There were no significant differences in biochemical recurrence in SPOPmut cancers. Limitations of this study include missing mutational data due to sample quality and lack of power to identify a difference in clinical outcomes. CONCLUSION: SPOP is mutated in 4.6% to 14.4% of patients with prostate cancer across different ethnic and demographic backgrounds. There was no significant association between SPOP mutations with ethnicity, clinical, or pathologic parameters. Mutual exclusivity of SPOP mutation with ERG rearrangement as well as a high association with CHD1 deletion reinforces SPOP mutation as defining a distinct molecular subclass of prostate cancer.

Candidate cell and matrix interaction domains on the collagen fibril, the predominant protein of vertebrates.

Type I collagen, the predominant protein of vertebrates, polymerizes with type III and V collagens and non-collagenous molecules into large cable-like fibrils, yet how the fibril interacts with cells and other binding partners remains poorly understood. To help reveal insights into the collagen structure-function relationship, a data base was assembled including hundreds of type I collagen ligand binding sites and mutations on a two-dimensional model of the fibril. Visual examination of the distribution of functional sites, and statistical analysis of mutation distributions on the fibril suggest it is organized into two domains. The "cell interaction domain" is proposed to regulate dynamic aspects of collagen biology, including integrin-mediated cell interactions and fibril remodeling. The "matrix interaction domain" may assume a structural role, mediating collagen cross-linking, proteoglycan interactions, and tissue mineralization. Molecular modeling was used to superimpose the positions of functional sites and mutations from the two-dimensional fibril map onto a three-dimensional x-ray diffraction structure of the collagen microfibril in situ, indicating the existence of domains in the native fibril. Sequence searches revealed that major fibril domain elements are conserved in type I collagens through evolution and in the type II/XI collagen fibril predominant in cartilage. Moreover, the fibril domain model provides potential insights into the genotype-phenotype relationship for several classes of human connective tissue diseases, mechanisms of integrin clustering by fibrils, the polarity of fibril assembly, heterotypic fibril function, and connective tissue pathology in diabetes and aging.