Preliminary investigation of crosslinked chitosan sponges for tailorable drug delivery and infection control

Authors

Ashley C. Parker, Jessica A. Jennings, Joel D. Bumgardner, Harry S. Courtney, Ernö Lindner, Warren O. Haggard

Abstract

Local versus systemic antibiotic delivery may be an effective strategy for treating musculoskeletal infections, especially when antibiotic-resistant bacteria are present. Lyophilized uncrosslinked, genipin crosslinked, and genipin crosslinked with poly(N-isopropylacrylamide) (PNIPAM) chitosan sponges were analyzed for their in vitro degradation rate, chemical crosslinking, antibiotic uptake, elution, biologic activity, and cytotoxicity. These evaluations were pursued to determine if crosslinking with genipin could be used to create a tailorable point of care loaded sponge for local infection control. Crosslinking the chitosan sponges decreased degradation in phosphate-buffered saline from 4.48 ± 2.28 wt % remaining of the uncrosslinked sponges to 78.82 ± 1.15 and 73.87 ± 1.27 wt % remaining at week 1 for the genipin and PNIPAM/genipin crosslinked sponges, respectively. The PNIPAM/genipin crosslinked sponges exhibited the most sustained release of biologically active antibiotics, with an average antibiotic release 63% higher than uncrosslinked and 37% higher than genipin crosslinked sponges, after 96 h. No significant cytotoxic effects from sponges or eluates were exhibited with NIH 3T3 fibroblasts. These preliminary results indicate that genipin crosslinked chitosan sponges, with or without PNIPAM, have potential as local delivery systems for adjunctive therapy for infection control, especially when longer degradation periods and higher antibiotic elutions are desired.

Link to Articles

http://dx.doi.org/10.1002/jbm.b.32822

Halofuginone inhibits the establishment and progression of melanoma bone metastases

Authors

Patricia Juárez, Khalid S. Mohammad, Juan Juan Yin, Pierrick G. J. Fournier, Ryan C. McKenna, Holly W Davis, Xiang H. Peng, Maria Niewolna, Delphine Javelaud, John M Chirgwin, Alain Mauviel, and Theresa A. Guise

Abstract

Transforming growth factor (TGF-β) derived from bone fuels melanoma bone metastases by inducing tumor secretion of pro-metastatic factors that act on bone cells to change the skeletal microenvironment. Halofuginone is a plant alkaloid derivative that blocks TGF-β signaling with antiangiogenic and antiproliferative properties. Here, we demonstrate for the first time that halofuginone therapy decreases development and progression of bone metastasis caused by melanoma cells through inhibition of TGF-β signaling. Halofuginone treatment of human melanoma cells inhibited cell proliferation, phosphorylation of SMAD proteins in response to TGF-β, and TGF-β-induced SMAD-driven transcription. In addition, halofuginone reduced expression of TGF-β target genes that enhance bone metastases, including PTHrP, CTGF, CXCR4, and IL11. Also, cell apoptosis was increased in response to halofuginone. In nude mice inoculated with 1205Lu melanoma cells, a preventive protocol with halofuginone inhibited bone metastasis. The beneficial effects of halofuginone treatment were comparable to those observed with other anti-TGF-β strategies, including systemic administration of SD208, a small molecule inhibitor of TGF-β receptor I kinase, or forced overexpression of Smad7, a negative regulator of TGF-β signaling. Furthermore, mice with established bone metastases treated with halofuginone had significantly less osteolysis than mice receiving placebo assessed by radiographys. Thus, halofuginone is also effective in reducing the progression of melanoma bone metastases. Moreover, halofuginone treatment reduced melanoma metastasis to the brain, showing the potential of this novel treatment against cancer metastasis.

Link to Article

http://dx.doi.org/10.1158/0008-5472.CAN-12-1444

Creation and Preliminary Characterization of a Leptin Knockout Rat

Authors

Sergio Vaira, Chang Yang, Aaron McCoy, Kelly Keys, Shurong Xue, Edward J. Weinstein, Deborah V. Novack and Xiaoxia Cui

Abstract

Leptin, a cytokine-like hormone secreted mainly by adipocytes, regulates various pathways centered on food intake and energy expenditure, including insulin sensitivity, fertility, immune system, and bone metabolism. Here, using zinc finger nuclease technology, we created the first leptin knockout rat. Homozygous leptin null rats are obese with significantly higher serum cholesterol, triglyceride, and insulin levels than wild-type controls. Neither gender produced offspring despite of repeated attempts. The leptin knockout rats also have depressed immune system. In addition, examination by microcomputed tomography of the femurs of the leptin null rats shows a significant increase in both trabecular bone mineral density and bone volume of the femur compared with wild-type littermates. Our model should be useful for many different fields of studies, such as obesity, diabetes, and bone metabolism-related illnesses.

Link to Article

http://dx.doi.org/10.1210/en.2012-1462

RANKL inhibition combined with tamoxifen treatment increases anti-tumor efficacy and prevents tumor-induced bone destruction in an estrogen receptor-positive breast cancer bone metastasis model

Authors

Jude Canon, Rebecca Bryant, Martine Roudier, Daniel G. Branstetter and William C. Dougall

Abstract

Tumor cells in bone can induce the activation of osteoclasts, which mediate bone resorption and release of growth factors and calcium from the bone matrix, resulting in a cycle of tumor growth and bone breakdown. Targeting the bone microenvironment by the inhibition of RANKL, an essential mediator of osteoclast function, not only prevents tumor-induced osteolysis but also decreases skeletal tumor burden in preclinical models. The inhibition of skeletal tumor progression after the inhibition of osteoclasts is via interruption of the “vicious cycle” of tumor/bone interactions. The majority of breast cancer patients at risk for bone metastases harbor estrogen receptor-positive (ER+) tumors. We developed a mouse model for ER+ breast cancer bone metastasis and evaluated the effect of RANKL inhibition on tumor-induced osteolysis and skeletal tumor growth both alone and in combination with tamoxifen. Luciferase-labeled MCF-7 cells (MCF-7Luc) formed metastatic foci in the hind limbs following intracardiac injection and caused mixed osteolytic/osteoblastic lesions. RANKL inhibition by OPG-Fc treatment blocked osteoclast activity and prevented tumor-induced osteolysis, as well as caused a marked decrease in skeletal tumor burden. Tamoxifen as a single agent reduced MCF-7Luc tumor growth in the hind limbs. In a combination experiment, OPG-Fc plus tamoxifen resulted in significantly greater tumor growth inhibition than either single agent alone. Histologic analysis revealed a decrease in the proliferation of tumor cells by both single agents, which was enhanced in the combination treatment. Upon treatment with OPG-Fc alone or in combination with tamoxifen, there was a complete absence of osteolytic lesions, demonstrating the ability of RANKL inhibition to prevent skeletal related morbidity in an ER+ model. The combination approach of targeting osteoclasts and the bone microenvironment by RANKL inhibition and the tumor directly via hormonal therapy may provide additional benefit to reducing skeletal tumor progression in ER+ breast cancer patients.

Link to Article

http://dx.doi.org/10.1007/s10549-012-2222-2

Effects of androgen deprivation therapy and bisphosphonate treatment on bone in patients with...

Authors

Colm Morrissey, Martine P. Roudier, Alex Dowell, Lawrence D. True, Melanie Ketchanji, Christopher Welty, Eva Corey, Paul H. Lange, Celestia S. Higano, Robert L. Vessella

Abstract

Qualitative and quantitative bone features were determined in nondecalcified and decalcified bone from 20 predetermined bone sites in each of 44 patients who died with castration resistant prostate cancer (CRPC), some of which received bisphosphonate treatment (BP) in addition to androgen deprivation therapy (ADT). Thirty nine of the 44 patients (89%) had evidence of bone metastases. By histomorphometric analysis, these bone metastases were associated with a range of bone responses from osteoblastic to osteolytic with a wide spectrum of bone responses often seen within an individual patient. Overall, the average bone volume/tissue volume (BV/TV) was 25.7% confirming the characteristic association of an osteoblastic response to prostate cancer bone metastasis when compared to the normal age-matched weighted mean BV/TV of 14.7%. The observed new bone formation was essentially woven bone and this was a localized event. In comparing BV/TV at metastatic sites between patients who had received BP treatment and those that had not, there was a significant difference (28.6% vs 19.3%, respectively). At bone sites that were not invaded by tumor, the average BV/TV was 10.1% indicating significant bone loss due to ADT that was not improved (11%) in those patients who had received BPs. Surprisingly there was no significant difference in the number of osteoclasts present at the metastatic sites between patients treated or not treated with BPs but in bone sites where the patient had been treated with BPs, giant osteoclasts were observed. Overall, 873 paraffin embedded specimens and 661 methylmethacrylate embedded specimens were analyzed. Our results indicate that in CRPC patients, ADT induces serious bone loss even in patients treated with BP. Furthermore, in this cohort of patients, BP treatment increased BV and did not decrease the number of osteoclasts in prostate cancer bone metastases compared to bone metastases from patients who did not receive BP.

Link to Article

http://dx.doi.org/10.1002/jbmr.1749

Mesenchymal stem cells increase collagen infiltration and improve wound healing response to porous titanium percutaneous implants

Authors

Dorthyann Isackson, Kevin J. Cook, Lawrence D. McGill, Kent N. Bachus

Abstract

Epidermal downgrowth, commonly associated with long-term percutaneous implants, weakens the skin-implant seal and greatly increases the vulnerability of the site to infection. To improve the skin attachment and early tissue integration with porous metal percutaneous implants, we evaluated the effect of bone marrow-derived mesenchymal stem cells (BMMSCs) to provide wound healing cues and vascularization to the dermal and epidermal tissues in establishing a barrier with the implant. Two porous metal percutaneous implants, one treated with BMMSCs and one untreated, were placed subdermally on the dorsum of Lewis rats. Implants were evaluated at 0, 3, 7, 28, and 56 days after implantation. Histological analyses evaluated cellular infiltrates, vascularization, quantity and quality of tissue ingrowth, epidermal downgrowth, and fibrous encapsulation. The amount of collagen infiltrating the porous coating was significantly greater for the BMMSC-treated implants at 3 and 28 days post implantation compared to untreated implants. There was an early influx and resolution of cellular inflammatory infiltrates in the treated implants compared to the untreated, though not statistically significant. Vascularization increased over time in both treated and untreated implants, with no statistical significance. Epidermal downgrowth was minimally observed in all implants with or without the BMMSC treatment. Our results suggest that BMMSCs can influence an early and rapid resolution of acute and chronic inflammation in wound healing, and can stimulate early collagen deposition and granulation tissue associated with later stages of wound repair. These findings provide evidence that BMMSCs can stimulate a more rapid and improved barrier between the skin and porous metal percutaneous implant.

Link to Article

http://dx.doi.org/10.1016/j.medengphy.2012.08.002