Development of Gendine-Coated Cannula for Continuous Subcutaneous Insulin Infusion for Extended Use

Continuous subcutaneous insulin infusion (CSII) using pumps is a widely used method for insulin therapy in patients with diabetes mellitus. Among the major factors that usually lead to the discontinuation of CSII are CSII set-related issues, including infection at the infusion site. The American Diabetic Association currently recommends rotating sites every 2 to 3 days. This recommendation adds cost and creates inconvenience. Therefore, in order to prevent infections and extend the duration between insertion site changes, we developed a Teflon cannula coated with a combination of gentian violet and chlorhexidine (gendine) and tested its antimicrobial efficacy against different pathogens. The cannulas were coated with gendine on the exterior surface and dried. The efficacy and durability of gendine-coated cannulas were determined against methicillin-resistant Staphylococcus aureus, Staphylococcus epidermidis, methicillin-susceptible S. aureus, Streptococcus pyogenes, vancomycin-resistant enterococci, Escherichia coli, Pseudomonas aeruginosa, Candida albicans, and Candida glabrata using a biofilm colonization method. The cytotoxicity of gendine was assessed against mouse fibroblast cell lines. The gendine-coated cannulas showed complete prevention of biofilm colonization of all organisms tested for up to 2 weeks (P < 0.0001) compared to that with the uncoated control. A gendine-coated catheter against mouse fibroblast cells was shown to be noncytotoxic. Our in vitro results show that a novel gendine cannula is highly effective in completely inhibiting the biofilm of multidrug-resistant pathogens for up to 2 weeks and may have potential clinical applications, such as prolonged use, cost reduction, and lower infection rate.     

Contrast-induced nephropathy after percutaneous coronary interventions in relation to chronic kidney disease and hemodynamic variables

We previously found that contrast-induced nephropathy (CIN) complicating percutaneous coronary intervention adversely affects patients with chronic kidney disease (CKD). Therefore, we further investigated whether the predictors and outcome of CIN after percutaneous coronary intervention differ among patients with versus without CKD. Among 7,230 consecutive patients, CIN (>or=25% or >or=0.5 mg/dl increase in preprocedure serum creatinine 48 hours after the procedure) developed in 381 of 1,980 patients (19.2%) with baseline CKD (estimated glomerular filtration rate [eGFR] <60 ml/min/1.73 m(2)) and in 688 of 5,250 patients (13.1%) without CKD. Decreased eGFRs, periprocedural hypotension, higher contrast media volumes, lower baseline hematocrit, diabetes, pulmonary edema at presentation, intra-aortic balloon pump use, and ejection fraction <40% were the most significant predictors of CIN in patients with CKD. Apart from intra-aortic balloon pump use, predictors of CIN in patients without CKD were the same as mentioned, plus older age and type of contrast media. Regardless of baseline renal function, CIN correlated with longer in-hospital stay and higher rates of in-hospital complications and 1-year mortality compared with patients without CIN. By multivariate analysis, CIN was 1 of the most powerful predictors of 1-year mortality in patients with preexisting CKD (odds ratio 2.37, 95% confidence interval 1.63 to 3.44) or preserved eGFR (odds ratio 1.78; 95% confidence interval 1.22 to 2.60). Thus, regardless of the presence of CKD, baseline characteristics and periprocedural hemodynamic parameters predict CIN, and this complication is associated with worse in-hospital and 1-year outcomes.     

Mutation in Osteoactivin Decreases Bone Formation in Vivo and Osteoblast Differentiation in Vitro

We have previously identified osteoactivin (OA), encoded by Gpnmb, as an osteogenic factor that stimulates osteoblast differentiation in vitro. To elucidate the importance of OA in osteogenesis, we characterized the skeletal phenotype of a mouse model, DBA/2J (D2J) with a loss-of-function mutation in Gpnmb. Microtomography of D2J mice showed decreased trabecular mass, compared to that in wild-type mice [DBA/2J-Gpnmb⁺/SjJ (D2J/Gpnmb⁺)]. Serum analysis showed decreases in OA and the bone-formation markers alkaline phosphatase and osteocalcin in D2J mice. Although D2J mice showed decreased osteoid and mineralization surfaces, their osteoblasts were increased in number, compared to D2J/Gpnmb⁺ mice. We then examined the ability of D2J osteoblasts to differentiate in culture, where their differentiation and function were decreased, as evidenced by low alkaline phosphatase activity and matrix mineralization. Quantitative RT-PCR analyses confirmed the decreased expression of differentiation markers in D2J osteoblasts. In vitro, D2J osteoblasts proliferated and survived significantly less, compared to D2J/Gpnmb⁺ osteoblasts. Next, we investigated whether mutant OA protein induces endoplasmic reticulum stress in D2J osteoblasts. Neither endoplasmic reticulum stress markers nor endoplasmic reticulum ultrastructure were altered in D2J osteoblasts. Finally, we assessed underlying mechanisms that might alter proliferation of D2J osteoblasts. Interestingly, TGF-β receptors and Smad-2/3 phosphorylation were up-regulated in D2J osteoblasts, suggesting that OA contributes to TGF-β signaling. These data confirm the anabolic role of OA in postnatal bone formation.Osteoporosis is a growing public health problem, in part because of the increasing numbers of people living beyond the age of 65 years.¹ It is characterized by low bone mass due to increased bone resorption by osteoclasts and decreased bone formation by osteoblasts, with significant deterioration in the bone microarchitecture leading to high bone fragility and increased fracture risk.^1,2 The net effect of osteoporosis is low bone mass.¹ There is an increasing demand for identifying novel bone anabolic factors with potential therapeutic benefits in treating generalized bone loss, such as osteoporosis and/or major skeletal fracture.Osteoactivin is a novel glycoprotein first identified in natural mutant osteopetrotic rats.³ The same protein has been identified and named separately in several other species: as dendritic cell heparan sulfate proteoglycan integrin dependent ligand (DCHIL) in mouse dendritic cells,⁴ as transmembrane glycoprotein NMB (GPNMB) in human melanoma cell lines and melanocytes,⁵ and as hematopoietic growth factor inducible neurokinin (HGFIN) in human tumor cells.⁶ The current recommended name for the protein encoded by Gpnmb in mouse is transmembrane glycoprotein NMB (http://www.ncbi.nlm.nih.gov/protein/Q99P91.2); here, we continue to use osteoactivin (OA) for the protein and Gpnmb for the gene. OA is a type I transmembrane protein that consists of multiple domains, including an extracellular domain, transmembrane domain, and protein sorting signal sequence.⁷ Within the C-terminal domain, OA has an RGD motif, predicting an integrin attachment site.^3,7–9Our research group initially reported on the novel role of OA in osteoblast differentiation and function.^7–10 We demonstrated that OA expression has a temporal pattern during osteoblast differentiation, being highest during matrix maturation and culture mineralization in vitro.^7–11 Using loss-of–function and gain-of–function approaches in osteoblasts, we reported that OA overexpression increases osteoblast differentiation and function and that OA down-regulation decreases nodule formation, alkaline phosphatase (ALP) activity, osteocalcin (OC) production, and matrix mineralization in vitro.⁷ We also reported on the positive role of OA in mesenchymal stem cell (MSCs) differentiation into osteoblasts in vitro.¹² In another study, we showed that recombinant OA protein induces higher osteogenic potential of fetal-derived MSCs, compared with bone marrow–derived MSCs¹³ and its osteogenic effects in the mouse C3H10T1/2 MSC cell line were similar to those of recombinant BMP-2.¹² We also localized OA protein as associated predominately with osteoblasts lining trabecular bones in vivo,¹¹ and showed that local injection of recombinant OA increased bone mass in a rat model.¹⁴ Moreover, in a fracture repair model OA expression increased over time, reaching a maximum 2 weeks after fracture.¹¹ In a parallel study, recombinant OA supported bone regeneration and formation in a rat critical-size calvarial defect model.¹⁵ Others have shown that OA is highly expressed by osteoclasts in vitro, suggesting that it may regulate osteoclast formation and activity.¹⁶There is urgent need for an animal model to fully examine the role of OA in osteogenesis. Interestingly, a natural mutation of the Gpnmb gene has been identified in the DBA/2J (D2J) mouse strain.¹⁷ These mice exhibit high-frequency hearing loss, which begins at the time of weaning and becomes severe by 2 to 3 months of age.^18,19 Aged D2J mice also develop progressive eye abnormalities that closely mimic human hereditary glaucoma. The onset of disease symptoms falls roughly between 3 and 4 months of age, and disease becomes severe by 6 months of age.^5,20 D2J mice are homozygous for a nonsense mutation in the Gpnmb gene sequence that induces an early stop codon, generating a truncated protein sequence of 150 amino acids (aa) instead of the full-length 562-aa OA protein.⁵ The control for the D2J mouse is the wild-type DBA/2J-Gpnmb⁺/SjJ mouse (D2J/Gpnmb⁺), homozygous for the wild-type Gpnmb gene.²¹ These Gpnmb wild-type mice do not develop glaucoma, as D2J mice do, although they exhibit mild iris stromal atrophy.²¹In the present study, we used Gpnmb mutant (D2J) and Gpnmb wild-type (D2J/Gpnmb⁺) mice to gain insight into the role of OA in osteogenesis and in osteoblast differentiation and function. Here, we report that loss-of–function mutation of Gpnmb suppresses bone formation by directly affecting osteoblast proliferation and survival, leading to a decreased number of differentiated osteoblasts with suppressed activity in bone mineralization. Thus, our data point to OA as a novel and positive regulator of postnatal bone formation.     

Optimization of the Esperanza window trap for the collection of the African onchocerciasis vector Simulium damnosum sensu lato

A simple inexpensive trap (Esperanza window trap) was shown recently to collect significant numbers of Simulium ochraceum sensu lato, a major vector of Onchocerca volvulus in Mesoamerica. Here, we report studies optimizing this trap for the collection of Simulium damnosum s.l., the major vector of O. volvulus in Africa. A shortened, blue and black striped version of the Esperanza window trap, when baited with a combination of CO₂ and worn trousers, rivalled human landing collections in the number of S. damnosum s.l. females collected. Traps baited with a commercially available human skin lure and CO₂ resulted in collections that were not significantly different than those obtained from traps baited with worn trousers and CO₂. This suggests that the Esperanza window trap may offer a replacement for human landing collections for monitoring onchocerciasis transmission in Africa.