Faculty Bibliography

Receptor activator of NF-B ligand (RANKL) stimulation leads to the activation of mitogen-activated protein kinase (MAPK)/AP-1 and Ca ²⁺-nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) signaling pathways in osteoclastogenesis. Targeting these pathways has been an encouraging strategy for bone-related diseases, such as postmenopausal osteoporosis. In this study, we examined the effects of caffeic acid 3,4-dihydroxy-phenethyl ester (CADPE) on osteoclastogenesis. In mouse bone marrow monocytes (BMMs) and RAW264.7 cells, CADPE suppressed RANKL-induced osteoclast differentiation and actin-ring formation in a dose-dependent manner within non-growth inhibitory concentrations at the early stage, while CADPE had no effect on macrophage colony-stimulating factor (M-CSF)-induced proliferation and differentiation. At the molecular level, CADPE inhibited RANKL-induced phosphorylation of MAPKs, including extracellular signal-regulated kinases 1/2 (ERK1/2), p38, and c-Jun N-terminal kinase (JNK), without significantly affecting the NF-B signaling pathway. CADPE abrogated RANKL-induced activator protein 1 (AP-1)/FBJ murine osteosarcoma viral oncogene homolog (c-Fos) nuclear translocation and activation. Overexpression of c-Fos prevented the inhibition by CADPE of osteoclast differentiation. Furthermore, CADPE suppressed RANKL-induced the tumor necrosis factor receptor associated factor 6 (TRAF6) interaction with c-src tyrosine kinase (c-Src), blocked RANKL-induced the phosphorylation of protein kinase B (AKT), and inhibited RANKL-induced Ca²⁺ oscillation. As a result, CADPE decreased osteoclastogenesis- related marker gene expression, including NFATc1, TRAP, cathepsin K, and c-Src. To test the effects of CADPE on osteoclast activity in vivo, we showed that CADPE prevented ovariectomy-induced bone loss by inhibiting osteoclast activity. Together, our data demonstrate that CADPE suppresses osteoclastogenesis and bone loss through inhibiting RANKL-induced MAPKs and Ca²⁺-NFATc1 signaling pathways. CADPE is a novel agent in the treatment of osteoclast-related diseases, such as osteoporosis

OBJECTIVES: The reported long-term safety of kidney donation is inconsistent with the impairment of kidney function observed following nephrectomy for renal cell cancer. We aimed to investigate if indication for nephrectomy (kidney cancer vs. living donation) was an independent risk factor for kidney function deterioration. MATERIALS AND METHODS: Between 1985 and 2008, 124 patients with localized renal cell carcinoma who meet the criteria used for living donation, underwent radical nephrectomy (group 1) at our institution. Group 1 was retrospectively compared with 124 consecutive living donor nephrectomies (group 2) performed from 2004 to 2008. Kidney function evaluation was performed preoperatively and at 1, 2, 3, and 4 years postoperatively with calculation of estimated glomerular filtration rate through the Modification of Diet in Renal Disease (MDRD-eGFR) and the adjusted Cockroft and Gault (CG-eGFR) formula. Multivariate logistic regression included patients' characteristics and indication for nephrectomy as predictors of kidney function deterioration. RESULTS: Mean decrease in MDRD-eGFR was 30.4% and 32.4% in groups 1 and 2 (P = 0.30). Prevalence of chronic kidney disease (CKD), defined by MDRD-eGFR < 60 mL/min/m(2), varied from 42.3% to 71% in group 1 and from 41.6% to 56% in group 2 at different time points (P = 0.073). Prevalence of CKD at 4 years defined by MDRD-eGFR < 45 mL/min/m(2) was significantly increased in group 1 compared with group 2 (16.2% and 5.3%, P < 0.005, respectively). Linear regression analysis showed only baseline kidney function and patient age predicted a significant decrease in postoperative kidney function (P < 0.001 and P = 0.04). CONCLUSIONS: Renal cell carcinoma is not an independent risk factor for kidney function impairment following nephrectomy. Selected kidney cancer patients with few morbidities face the same deterioration of meanly 30% of kidney function compared with living donors, but their lower baseline function results in an increased risk for CKD.

The basement membrane between the epidermis and the dermis is indispensable for normal skin functions. It connects, and functionally separates, the epidermis and the dermis. To understand the suprastructural and functional basis of these connections, heterotypic supramolecular aggregates were isolated from the dermal-epidermal junction zone of human skin. Individual suprastructures were separated and purified by immunomagnetic beads, each recognizing a specific, molecular component of the aggregates. The molecular compositions of the suprastructures were determined by immunogold electron microscopy and immunoblotting. A composite of two networks was obtained from fibril-free suspensions by immunobeads recognizing either laminin 332 or collagen IV. After removal of perlecan-containing suprastructures or after enzyme digestion of heparan sulfate chains, a distinct network with a diffuse electron-optical appearance was isolated with magnetic beads coated with antibodies to collagen IV. The second network was more finely grained and comprised laminin 332 and laminins with alpha5-chains. The core protein of perlecan was an exclusive component of this network whereas its heparan sulfate chains were integrated into the collagen IV-containing network. Nidogens 1 and 2 occurred in both networks but did not form strong molecular cross-bridges. Their incorporation into one network appeared to be masked after their incorporation into the other one. We conclude that the epidermal basement membrane is a composite of two structurally independent networks that are tightly connected in a spot-welding-like manner by perlecan-containing aggregates.

Prosthesis loosening, associated with wear particle-induced inflammation and osteoclast-mediated bone destruction, is a common cause for joint implant failure, leading to revision surgery. Adenosine A(2A) receptors (A(2A)Rs) mediate potent anti-inflammatory effects in many tissues and prevent osteoclast differentiation. We tested the hypothesis that an A(2A)R agonist could reduce osteoclast-mediated bone resorption in a murine calvaria model of wear particle-induced bone resorption. C57BL/6 and A(2A)R knockout (A(2A)R KO) mice received ultrahigh-molecular weight polyethylene particles and were treated daily with either saline or the A(2A)R agonist CGS21680. After 2 weeks, micro-computed tomography of calvaria demonstrated that CGS21680 reduced particle-induced bone pitting and porosity in a dose-dependent manner, increasing cortical bone and bone volume compared to control mice. Histological examination demonstrated diminished inflammation after treatment with CGS21680. In A(2A)R KO mice, CGS21680 did not affect osteoclast-mediated bone resorption or inflammation. Levels of bone resorption markers receptor activator of nuclear factor kappaB (RANK), RANK ligand, cathepsin K, CD163, and osteopontin were reduced after CGS21680 treatment, together with a reduction in osteoclasts. Secretion of interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha was significantly decreased, whereas IL-10 was markedly increased in bone by CGS21680. These results in mice suggest that site-specific delivery of an adenosine A(2A)R agonist could enhance implant survival, delaying or eliminating the need for revision arthroplastic surgery.

Physical damage to the peripheral nerves triggers Schwann cell injury response in the distal nerves in an event termed Wallerian degeneration: the Schwann cells degrade their myelin sheaths and dedifferentiate, reverting to a phenotype that supports axon regeneration and nerve repair. The molecular mechanisms regulating Schwann cell plasticity in the PNS remain to be elucidated. Using both in vivo and in vitro models for peripheral nerve injury, here we show that inhibition of p38 mitogen-activated protein kinase (MAPK) activity in mice blocks Schwann cell demyelination and dedifferentiation following nerve injury, suggesting that the kinase mediates the injury signal that triggers distal Schwann cell injury response. In myelinating cocultures, p38 MAPK also mediates myelin breakdown induced by Schwann cell growth factors, such as neuregulin and FGF-2. Furthermore, ectopic activation of p38 MAPK is sufficient to induce myelin breakdown and drives differentiated Schwann cells to acquire phenotypic features of immature Schwann cells. We also show that p38 MAPK concomitantly functions as a negative regulator of Schwann cell differentiation: enforced p38 MAPK activation blocks cAMP-induced expression of Krox 20 and myelin proteins, but induces expression of c-Jun. As expected of its role as a negative signal for myelination, inhibition of p38 MAPK in cocultures promotes myelin formation by increasing the number as well as the length of individual myelin segments. Altogether, our data identify p38 MAPK as an important regulator of Schwann cell plasticity and differentiation.

Establishment of germline sexual identity is critical for production of male and female germline stem cells, as well as sperm versus eggs. Here we identify PHD Finger Protein 7 (PHF7) as an important factor for male germline sexual identity in Drosophila. PHF7 exhibits male-specific expression in early germ cells, germline stem cells, and spermatogonia. It is important for germline stem cell maintenance and gametogenesis in males, whereas ectopic expression in female germ cells ablates the germline. Strikingly, expression of PHF7 promotes spermatogenesis in XX germ cells when they are present in a male soma. PHF7 homologs are also specifically expressed in the mammalian testis, and human PHF7 rescues Drosophila Phf7 mutants. PHF7 associates with chromatin, and both the human and fly proteins bind histone H3 N-terminal tails with a preference for dimethyl lysine 4 (H3K4me2). We propose that PHF7 acts as a conserved epigenetic "reader" that activates the male germline sexual program.

Human influenza viruses are proposed to recognize sialic acids (pink diamonds) on glycans extended with poly-LacNAc chains (LacNAc=(yellow circle+blue square)). N- and O-linked glycans were extended with different poly-LacNAc chains with alpha2-3- and alpha2-6-linked sialic acids recognized by human and avian influenza viruses, respectively. The specificity of recombinant hemagglutinins (receptors in green) was investigated by using glycan microarray technology.