A Heparan Sulfate-Binding Cell Penetrating Peptide for Tumor Targeting and Migration Inhibition.

As heparan sulfate proteoglycans (HSPGs) are known as co-receptors to interact with numerous growth factors and then modulate downstream biological activities, overexpression of HS/HSPG on cell surface acts as an increasingly reliable prognostic factor in tumor progression. Cell penetrating peptides (CPPs) are short-chain peptides developed as functionalized vectors for delivery approaches of impermeable agents. On cell surface negatively charged HS provides the initial attachment of basic CPPs by electrostatic interaction, leading to multiple cellular effects. Here a functional peptide (CPPecp) has been identified from critical HS binding region in hRNase3, a unique RNase family member with in vitro antitumor activity. In this study we analyze a set of HS-binding CPPs derived from natural proteins including CPPecp. In addition to cellular binding and internalization, CPPecp demonstrated multiple functions including strong binding activity to tumor cell surface with higher HS expression, significant inhibitory effects on cancer cell migration, and suppression of angiogenesis in vitro and in vivo. Moreover, different from conventional highly basic CPPs, CPPecp facilitated magnetic nanoparticle to selectively target tumor site in vivo. Therefore, CPPecp could engage its capacity to be developed as biomaterials for diagnostic imaging agent, therapeutic supplement, or functionalized vector for drug delivery.

Low-level laser therapy alleviates neuropathic pain and promotes function recovery in rats with chronic constriction injury: possible involvements in hypoxia-inducible factor 1α (HIF-1α).

Nerve inflammation plays an important role in the development and progression of neuropathic pain after chronic constrictive injury (CCI). Recent studies have indicated that hypoxia-inducible factor 1α (HIF-1α) is crucial in inflammation. Low-level laser therapy has been used in treating musculoskeletal pain, but rare data directly support its use for neuropathic pain. We investigated the effects of low-level laser on the accumulation of HIF-1α, tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß) in controlling neuropathic pain, as well as on the activation of vascular endothelial growth factor (VEGF) and nerve growth factor (NGF) in promoting functional recovery in a rat CCI model. CCI was induced by placing four loose ligatures around the sciatic nerve of rats. Treatments of low-level laser (660 nm, 9 J/cm(2)) or sham irradiation (0 J/cm(2)) were performed at the CCI sites for 7 consecutive days. The effects of laser in animals with CCI were determined by measuring the mechanical paw withdrawal threshold, as well as the sciatic, tibial, and peroneal function indices. Histopathological and immunoassay analyses were also performed. Low-level laser therapy significantly improved paw withdrawal threshold and the sciatic, tibial, and peroneal functional indices after CCI. The therapy also significantly reduced the overexpressions of HIF-1α, TNF-α, and IL-1ß, and increased the amounts of VEGF, NGF, and S100 proteins. In conclusion, a low-level laser could modulate HIF-1α activity. Moreover, it may also be used as a novel and clinically applicable therapeutic approach for the improvement of tissue hypoxia/ischemia and inflammation in nerve entrapment neuropathy, as well as for the promotion of nerve regeneration. These findings might lead to a sufficient morphological and functional recovery of the peripheral nerve.

Killed but metabolically active Leishmania infantum as a novel whole-cell vaccine for visceral leishmaniasis.

There are currently no effective vaccines for visceral leishmaniasis, the second most deadly parasitic infection in the world. Here, we describe a novel whole-cell vaccine approach using Leishmania infantum chagasi promastigotes treated with the psoralen compound amotosalen (S-59) and low doses of UV A radiation. This treatment generates permanent, covalent DNA cross-links within parasites and results in Leishmania organisms termed killed but metabolically active (KBMA). In this report, we characterize the in vitro growth characteristics of both KBMA L. major and KBMA L. infantum chagasi. Concentrations of S-59 that generate optimally attenuated parasites were identified. Like live L. infantum chagasi, KBMA L. infantum chagasi parasites were able to initially enter liver cells in vivo after intravenous infection. However, whereas live L. infantum chagasi infection leads to hepatosplenomegaly in mice after 6 months, KBMA L. infantum chagasi parasites were undetectable in the organs of mice at this time point. In vitro, KBMA L. infantum chagasi retained the ability to enter macrophages and induce nitric oxide production. These characteristics of KBMA L. infantum chagasi correlated with the ability to prophylactically protect mice via subcutaneous vaccination at levels similar to vaccination with live, virulent organisms. Splenocytes from mice vaccinated with either live L. infantum chagasi or KBMA L. infantum chagasi displayed similar cytokine patterns in vitro. These results suggest that KBMA technology is a potentially safe and effective novel vaccine strategy against the intracellular protozoan L. infantum chagasi. This approach may represent a new method for whole-cell vaccination against other complex intracellular pathogens.

Immunomodulation by imiquimod in patients with high-risk primary melanoma.

Imiquimod is a synthetic Toll-like receptor 7 (TLR7) agonist approved for the topical treatment of actinic keratoses, superficial basal cell carcinoma, and genital warts. Imiquimod leads to an 80-100% cure rate of lentigo maligna; however, studies of invasive melanoma are lacking. We conducted a pilot study to characterize the local, regional, and systemic immune responses induced by imiquimod in patients with high-risk melanoma. After treatment of the primary melanoma biopsy site with placebo or imiquimod cream, we measured immune responses in the treated skin, sentinel lymph nodes (SLNs), and peripheral blood. Treatment of primary melanomas with 5% imiquimod cream was associated with an increase in both CD4+ and CD8+ T cells in the skin, and CD4+ T cells in the SLN. Most of the CD8+ T cells in the skin were CD25 negative. We could not detect any increases in CD8+ T cells specifically recognizing HLA-A(*)0201-restricted melanoma epitopes in the peripheral blood. The findings from this small pilot study demonstrate that topical imiquimod treatment results in enhanced local and regional T-cell numbers in both the skin and SLN. Further research into TLR7 immunomodulating pathways as a basis for effective immunotherapy against melanoma in conjunction with surgery is warranted.

Hyaluronan modulates accumulation of hypoxia-inducible factor-1 alpha, inducible nitric oxide synthase, and matrix metalloproteinase-3 in the synovium of rat adjuvant-induced arthritis model.

INTRODUCTION: Hypoxia is a feature of the inflamed synovium in rheumatoid arthritis (RA). Intra-articular injection of hyaluronan (HA) may be considered a potential way to treat RA. However, the exact molecular mechanism of HA on decreased cellular responses to hypoxic environment is unclear. The present study has been designed to use the adjuvant-induced arthritis model to examine the effects of HA on the changes of immunohistochemical expressions of hypoxia-inducible factor-1alpha (HIF-1alpha), inducible nitric oxide synthase (iNOS), and matrix metalloproteinase-3 (MMP3) in the synovial tissues at the early phase of arthritic inflammation. METHODS: Monoarthritis was induced in adult male Sprague-Dawley (250-300 g) via intraarticular injection of complete Freund's adjuvant (CFA) into the tibiotarsal joint. The CFA-induction arthritis animals were divided into three groups: treatment (intraarticular injection of HA), placebo (intraarticular injection of saline) and controls (no treatments). Functional evaluations of edema and pain behavior, histology, and HIF-1alpha, iNOS, and MMP3 immunohistochemistry were performed before, after the first injection, three injections, and on the follow-up injection of the treatments. RESULTS: Intra-articular injection of HA also significantly suppressed the mechanical allodynia (p < 0.001) and overexpressions of HIF-1alpha (p < 0.001), iNOS (p = 0.004) and MMP3 (p < 0.001) immunoreactivity in synovium. CONCLUSIONS: This study demonstrated that early intervention of HA is an effective protection against accumulation of inflammation-induced HIF-1alpha, iNOS, and MMP3 to limit erosive damage in CFA-induced model of arthritis.

Assessment of recoverable forms of sulfur particles used in bioleaching of contaminated sediments.

The use of recoverable sulfur particles will enhance the feasibility and reduce the cost of bioleaching process. Three different forms of sulfur particles, powder, pastilles and pellets were used to study the utilization and recovery of sulfur, used as energy source for thiobacilli in the bioleaching process. The Langmuir isotherm was used to explain the adsorption equilibrium existing between the sorbed and suspended bacteria and the maximum adsorption capacity obtained from the Langmuir isotherm was utilized to determine the specific surface area of the sulfur particles. The specific surface area of sulfur particles was found to be the determining factor in the bioleaching process and not the particle size. The rates of pH reduction, sulfate production and metal solubilization increased with increasing specific surface area of the particles. The pH reduction and metal solubilization were significantly enhanced by the reuse of recovered sulfur particles. The efficiency of metal solubilization with recovered sulfur pastilles was comparable to that with sulfur powder. This study revealed the practicability of reusing the recovered sulfur pastilles in the bioleaching process.