Limits...
Identification of beta-2 as a key cell adhesion molecule in PCa cell neurotropic behavior: a novel ex vivo and biophysical approach.

Jansson KH, Castillo DG, Morris JW, Boggs ME, Czymmek KJ, Adams EL, Schramm LP, Sikes RA - PLoS ONE (2014)

Bottom Line: We overexpressed beta-2 as a fusion protein with enhanced cyan fluorescence protein (ECFP) in weakly aggressive LNCaP cells and observed neurotropic effects utilizing our novel ex vivo organotypic spinal cord co-culture model, and performed functional assays with neural matrices and atomic force microscopy.On laminin, a neural CAM, overexpression of beta-2 enhances PCa cell migration, invasion, and growth. 2BECFP cells exhibit marked binding affinity to laminin relative to LNECFP controls, and recombinant beta-2 ectodomain elicits more binding events to laminin than BSA control.Functional overexpression of VSSC beta subunits in PCa may mediate PCa metastatic behavior through association with neural matrices.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Cancer Ontogeny and Therapeutics, Department of Biological Sciences, University of Delaware, Newark, Delaware, United States of America; The Center for Translational Cancer Research, University of Delaware, Newark, Delaware, United States of America.

ABSTRACT
Prostate cancer (PCa) is believed to metastasize through the blood/lymphatics systems; however, PCa may utilize the extensive innervation of the prostate for glandular egress. The interaction of PCa and its nerve fibers is observed in 80% of PCa and is termed perineural invasion (PNI). PCa cells have been observed traveling through the endoneurium of nerves, although the underlying mechanisms have not been elucidated. Voltage sensitive sodium channels (VSSC) are multimeric transmembrane protein complexes comprised of a pore-forming α subunit and one or two auxiliary beta (β) subunits with inherent cell adhesion molecule (CAM) functions. The beta-2 isoform (gene SCN2B) interacts with several neural CAMs, while interacting putatively with other prominent neural CAMs. Furthermore, beta-2 exhibits elevated mRNA and protein levels in highly metastatic and castrate-resistant PCa. When overexpressed in weakly aggressive LNCaP cells (2BECFP), beta-2 alters LNCaP cell morphology and enhances LNCaP cell metastasis associated behavior in vitro. We hypothesize that PCa cells use beta-2 as a CAM during PNI and subsequent PCa metastasis. The objective of this study was to determine the effect of beta-2 expression on PCa cell neurotropic metastasis associated behavior. We overexpressed beta-2 as a fusion protein with enhanced cyan fluorescence protein (ECFP) in weakly aggressive LNCaP cells and observed neurotropic effects utilizing our novel ex vivo organotypic spinal cord co-culture model, and performed functional assays with neural matrices and atomic force microscopy. With increased beta-2 expression, PCa cells display a trend of enhanced association with nerve axons. On laminin, a neural CAM, overexpression of beta-2 enhances PCa cell migration, invasion, and growth. 2BECFP cells exhibit marked binding affinity to laminin relative to LNECFP controls, and recombinant beta-2 ectodomain elicits more binding events to laminin than BSA control. Functional overexpression of VSSC beta subunits in PCa may mediate PCa metastatic behavior through association with neural matrices.

Show MeSH

Related in: MedlinePlus

Overexpression of beta-2 enhances LNCaP cell migration, invasion, and growth on laminin.A. Quantified results of wound healing migration assays performed on 60 mm dishes pre-coated with 0.5 µg/mL laminin. 2BECFP cells exhibit a trend in increased migration on laminin compared to LNECFP cells beginning at day 1, with statistical significance achieved at days 3 and 4 (P<0.05, n = 5). B. LNCaP cells engineered to overexpress beta-2 have a 2.2 fold increase in invasion through 50 µg/ml laminin compared to vector control LNCaPs (P< 0.05, n = 3). C. Quantified results of MTT assays on 0.5 µg/mL laminin. 2BECFP cells have a statistically significant growth advantage on laminin relative to LNECFP cells beginning at day 5 (P< 0.05, n = 5).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4043823&req=5

pone-0098408-g004: Overexpression of beta-2 enhances LNCaP cell migration, invasion, and growth on laminin.A. Quantified results of wound healing migration assays performed on 60 mm dishes pre-coated with 0.5 µg/mL laminin. 2BECFP cells exhibit a trend in increased migration on laminin compared to LNECFP cells beginning at day 1, with statistical significance achieved at days 3 and 4 (P<0.05, n = 5). B. LNCaP cells engineered to overexpress beta-2 have a 2.2 fold increase in invasion through 50 µg/ml laminin compared to vector control LNCaPs (P< 0.05, n = 3). C. Quantified results of MTT assays on 0.5 µg/mL laminin. 2BECFP cells have a statistically significant growth advantage on laminin relative to LNECFP cells beginning at day 5 (P< 0.05, n = 5).

Mentions: Aggressive cancer cells have an enhanced ability to migrate and are typically highly motile. As previously demonstrated in scratch assays, beta-2 overexpression enhanced LNCaP cell migration on plastic tissue culture dishes [62]. We examined whether this behavior would be consistent on plates coated with 0.5 µg/mL laminin. A trend in increased migration of 2BECFP relative to LNECFP controls is observed at both 1 day and 2 days with statistical significance achieved at both 3 days and 4 days (Figure 4A, P<0.05).


Identification of beta-2 as a key cell adhesion molecule in PCa cell neurotropic behavior: a novel ex vivo and biophysical approach.

Jansson KH, Castillo DG, Morris JW, Boggs ME, Czymmek KJ, Adams EL, Schramm LP, Sikes RA - PLoS ONE (2014)

Overexpression of beta-2 enhances LNCaP cell migration, invasion, and growth on laminin.A. Quantified results of wound healing migration assays performed on 60 mm dishes pre-coated with 0.5 µg/mL laminin. 2BECFP cells exhibit a trend in increased migration on laminin compared to LNECFP cells beginning at day 1, with statistical significance achieved at days 3 and 4 (P<0.05, n = 5). B. LNCaP cells engineered to overexpress beta-2 have a 2.2 fold increase in invasion through 50 µg/ml laminin compared to vector control LNCaPs (P< 0.05, n = 3). C. Quantified results of MTT assays on 0.5 µg/mL laminin. 2BECFP cells have a statistically significant growth advantage on laminin relative to LNECFP cells beginning at day 5 (P< 0.05, n = 5).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4043823&req=5

pone-0098408-g004: Overexpression of beta-2 enhances LNCaP cell migration, invasion, and growth on laminin.A. Quantified results of wound healing migration assays performed on 60 mm dishes pre-coated with 0.5 µg/mL laminin. 2BECFP cells exhibit a trend in increased migration on laminin compared to LNECFP cells beginning at day 1, with statistical significance achieved at days 3 and 4 (P<0.05, n = 5). B. LNCaP cells engineered to overexpress beta-2 have a 2.2 fold increase in invasion through 50 µg/ml laminin compared to vector control LNCaPs (P< 0.05, n = 3). C. Quantified results of MTT assays on 0.5 µg/mL laminin. 2BECFP cells have a statistically significant growth advantage on laminin relative to LNECFP cells beginning at day 5 (P< 0.05, n = 5).
Mentions: Aggressive cancer cells have an enhanced ability to migrate and are typically highly motile. As previously demonstrated in scratch assays, beta-2 overexpression enhanced LNCaP cell migration on plastic tissue culture dishes [62]. We examined whether this behavior would be consistent on plates coated with 0.5 µg/mL laminin. A trend in increased migration of 2BECFP relative to LNECFP controls is observed at both 1 day and 2 days with statistical significance achieved at both 3 days and 4 days (Figure 4A, P<0.05).

Bottom Line: We overexpressed beta-2 as a fusion protein with enhanced cyan fluorescence protein (ECFP) in weakly aggressive LNCaP cells and observed neurotropic effects utilizing our novel ex vivo organotypic spinal cord co-culture model, and performed functional assays with neural matrices and atomic force microscopy.On laminin, a neural CAM, overexpression of beta-2 enhances PCa cell migration, invasion, and growth. 2BECFP cells exhibit marked binding affinity to laminin relative to LNECFP controls, and recombinant beta-2 ectodomain elicits more binding events to laminin than BSA control.Functional overexpression of VSSC beta subunits in PCa may mediate PCa metastatic behavior through association with neural matrices.

View Article: PubMed Central - PubMed

Affiliation: Laboratory for Cancer Ontogeny and Therapeutics, Department of Biological Sciences, University of Delaware, Newark, Delaware, United States of America; The Center for Translational Cancer Research, University of Delaware, Newark, Delaware, United States of America.

ABSTRACT
Prostate cancer (PCa) is believed to metastasize through the blood/lymphatics systems; however, PCa may utilize the extensive innervation of the prostate for glandular egress. The interaction of PCa and its nerve fibers is observed in 80% of PCa and is termed perineural invasion (PNI). PCa cells have been observed traveling through the endoneurium of nerves, although the underlying mechanisms have not been elucidated. Voltage sensitive sodium channels (VSSC) are multimeric transmembrane protein complexes comprised of a pore-forming α subunit and one or two auxiliary beta (β) subunits with inherent cell adhesion molecule (CAM) functions. The beta-2 isoform (gene SCN2B) interacts with several neural CAMs, while interacting putatively with other prominent neural CAMs. Furthermore, beta-2 exhibits elevated mRNA and protein levels in highly metastatic and castrate-resistant PCa. When overexpressed in weakly aggressive LNCaP cells (2BECFP), beta-2 alters LNCaP cell morphology and enhances LNCaP cell metastasis associated behavior in vitro. We hypothesize that PCa cells use beta-2 as a CAM during PNI and subsequent PCa metastasis. The objective of this study was to determine the effect of beta-2 expression on PCa cell neurotropic metastasis associated behavior. We overexpressed beta-2 as a fusion protein with enhanced cyan fluorescence protein (ECFP) in weakly aggressive LNCaP cells and observed neurotropic effects utilizing our novel ex vivo organotypic spinal cord co-culture model, and performed functional assays with neural matrices and atomic force microscopy. With increased beta-2 expression, PCa cells display a trend of enhanced association with nerve axons. On laminin, a neural CAM, overexpression of beta-2 enhances PCa cell migration, invasion, and growth. 2BECFP cells exhibit marked binding affinity to laminin relative to LNECFP controls, and recombinant beta-2 ectodomain elicits more binding events to laminin than BSA control. Functional overexpression of VSSC beta subunits in PCa may mediate PCa metastatic behavior through association with neural matrices.

Show MeSH
Related in: MedlinePlus