Shuji Isotani

Met-Independent Hepatocyte Growth Factor-mediated regulation of cell adhesion in human prostate cancer cells

BackgroundProstate cancer cells communicate reciprocally with the stromal cells surrounding them, inside the prostate, and after metastasis, within the bone. Each tissue secretes factors for interpretation by the other. One stromally-derived factor, Hepatocyte Growth Factor (HGF), was found twenty years ago to regulate invasion and growth of carcinoma cells. Working with the LNCaP prostate cancer progression model, we found that these cells could respond to HGF stimulation, even in the absence of Met, the only known HGF receptor. The new HGF binding partner we find on the cell surface may help to clarify conflicts in the past literature about Met expression and HGF response in cancer cells.MethodsWe searched for Met or any HGF binding partner on the cells of the PC3 and LNCaP prostate cancer cell models, using HGF immobilized on agarose beads. By using mass spectrometry analyses and sequencing we have identified nucleolin protein as a novel HGF binding partner. Antibodies against nucleolin (or HGF) were able to ameliorate the stimulatory effects of HGF on met-negative prostate cancer cells. Western blots, RT-PCR, and immunohistochemistry were used to assess nucleolin levels during prostate cancer progression in both LNCaP and PC3 models.ResultsWe have identified HGF as a major signaling component of prostate stromal-conditioned media (SCM) and have implicated the protein nucleolin in HGF signal reception by the LNCaP model prostate cancer cells. Antibodies that silence either HGF (in SCM) or nucleolin (on the cell surfaces) eliminate the adhesion-stimulatory effects of the SCM. Likewise, addition of purified HGF to control media mimics the action of SCM. C4-2, an LNCaP lineage-derived, androgen-independent human prostate cancer cell line, responds to HGF in a concentration-dependent manner by increasing its adhesion and reducing its migration on laminin substratum. These HGF effects are not due to shifts in the expression levels of laminin-binding integrins, nor can they be linked to expression of the known HGF receptor Met, as neither LNCaP nor clonally-derived C4-2 sub-line contain any detectable Met protein. Even in the absence of Met, small GTPases are activated, linking HGF stimulation to membrane protrusion and integrin activation. Membrane-localized nucelolin levels increase during cancer progression, as modeled by both the PC3 and LNCaP prostate cancer progression cell lines.ConclusionWe propose that cell surface localized nucleolin protein may function in these cells as a novel HGF receptor. Membrane localized nucleolin binds heparin-bound growth factors (including HGF) and appears upregulated during prostate cancer progression. Antibodies against nucleolin are able to ameliorate the stimulatory effects of HGF on met-negative prostate cancer cells. HGF-nucleolin interactions could be partially responsible for the complexity of HGF responses and met expression reported in the literature.

Soluble factors derived from stroma activated androgen receptor phosphorylation in human prostate LNCaP cells: roles of ERK/MAP kinase.

Accumulated evidence suggests stromal–epithelial interactions are critical to the progression of prostate cancer. In this study, we characterized AR phosphorylation in LNCaP cells co‐cultured with the conditioned medium (CM) from human prostate stromal fibroblasts.

Transforming growth factor‐β1 expression in early biopsy specimen predicts long‐term graft function following pediatric renal transplantation

The main cause of late graft loss or declining long‐term graft function is chronic allograft nephropathy (CAN), characterized by progressive interstitial fibrosis. Transforming growth factor (TGF)‐β1 plays a key role in fibrogenesis. We immunohistochemically investigated whether the degree of TGF‐β1 expression in early biopsy specimens routinely obtained from stable allografts at 100 d could predict fibrosis and graft dysfunction in the late phase. Patients were children with grafts from related donors. We immunohistochemically determined intracellular and extracellular expression of TGF‐β1 in the graft using LC antibody (LC) for intracellular TGF‐β1 and CC antibody (CC) for extracellular TGF‐β1. The change in creatinine clearance between 100 d and 3 yr after transplantation (ΔCcr) was used as an index of long‐term graft function. We also used image analysis to calculate the relative area involved by interstitial fibrosis in the trichrome‐stained section of graft biopsy specimens at 100 d and 3 yr, designating the change as ΔFI. ΔCcr was −4.2±9.4 mL/min in subjects with minimal early immunoreactivity for CC and −20.5±15.9 mL/min in subjects with strong reactivity (p<0.05). ΔCcr was −14.5±18.6 mL/min in subjects with minimal early immunoreactivity for LC and −11.7±12.8 mL/min in those with strong reactivity. ΔFI in subjects with minimal CC reactivity (1.28±4.11%) tended to be lower than that in subjects with strong reactivity (8.45±15.47%). Neither fibrosis at 100 d nor ΔFI differed between subjects with minimal and strong LC reactivity. Thus, strong extracellular TGF‐β1 expression in grafts at 100 d after transplantation is associated with a long‐term decline in graft function and tends to be associated with increased graft fibrosis at 3 yr.

V3-04 PRECISION SURGERY IN ROBOTIC PARTIAL NEPHRECTOMY

RESULTS: Robotic left nephrectomy and level II caval thrombectomy was performed successfully via a single-dock, supine approach. This method yielded excellent and early access to the IVC and left renal hilum, and allowed for concomitant nephrectomy/LND without re-positioning. Total operative time was 420 minutes with 330 minutes robotic console time (174 minutes for exposure, 27 minutes IVC clamp time, 84 minutes for nephrectomy/LND). EBL was 500cc without need for peri-operative transfusions and no intraoperative complications. Length of stay was 5 days and no major perioperative complications were noted. Outcomes compare favorably to previously reported robotic caval thrombectomy procedures employing the lateral approach. CONCLUSIONS: We demonstrate successful robotic left nephrectomy with Level II caval thrombectomy using a supine, singledock approach. To our knowledge, this is the first description of this approach for robotic caval thrombectomy. In appropriately selected patients, this versatile approach allows for rapid caval control, bilateral renal hilar access, and obviates the need for patient repositioning.