The success of the Edmonton Protocol for islet transplantation has provided new hope in the treatment of type 1 diabetes. This study reports on the assessment of 83 human islet grafts transplanted using the Edmonton Protocol since 1999. Cellular composition, as assessed by immunohistochemistry, showed a lower islet purity (ϳ40%) than has been reported in previous studies using dithizone staining to quantitate islet equivalents. Furthermore, grafts were found to contain substantial populations of exocrine and ductal tissue. Total cellular insulin transplanted was 8,097.6 ؎ 3,164.4 g/patient, and was significantly lower in bottom gradient layer grafts than top gradient layer or whole/combined grafts (P < 0.0005). A static incubation test for islet function gave a stimulation index of 3-4, although this measure did not correlate with posttransplant metabolic outcome. Furthermore, we confirmed a previously reported trend in which donor age affects islet yield and purity. It is important to note that a significant positive correlation was observed between the number of islet progenitor (ductal-epithelial) cells transplanted and long-term metabolic success as assessed an by intravenous glucose tolerance test at ϳ2 years posttransplant. In summary, careful assessment of islet graft composition is needed in a clinical transplantation program to accurately estimate islet purity and assess the contribution of other cell types present, such as islet progenitor cells.

BackgroundThe widespread problem of antibiotic resistance in pathogens such as Staphylococcus aureus has prompted the search for new antimicrobial approaches. In this study we report for the first time the use of a light-activated antimicrobial agent, methylene blue, to kill an epidemic methicillin-resistant Staphylococcus aureus (EMRSA-16) strain in two mouse wound models.ResultsFollowing irradiation of wounds with 360 J/cm2 of laser light (670 nm) in the presence of 100 μg/ml of methylene blue, a 25-fold reduction in the number of viable EMRSA was seen. This was independent of the increase in temperature of the wounds associated with the treatment. Histological examination of the wounds revealed no difference between the photodynamic therapy (PDT)-treated wounds and the untreated wounds, all of which showed the same degree of inflammatory infiltration at 24 hours.ConclusionThe results of this study demonstrate that PDT is effective at reducing the total number of viable EMRSA in a wound. This approach has promise as a means of treating wound infections caused by antibiotic-resistant microbes as well as for the elimination of such organisms from carriage sites.

Covalently coupling toluidine blue O-tiopronin to a gold nanoparticle forms an enhanced, exceptionally potent antimicrobial agent when activated by white light or 632 nm laser light. Aqueous solutions of tiopronin-gold nanoparticles had no antimicrobial effect and, when added to solutions of toluidine blue O, did not enhance the antimicrobial effect of the latter. The minimum bactericidal concentration of the covalently coupled toluidine blue O-tiopronin gold conjugate for Staphylococcus aureus was at least four times lower than that of free toluidine blue O.

aPDT treatment may augment periodontal treatment by increasing bacterial killing, inactivating bacterial virulence factors, and inactivating host cytokines that impair periodontal restoration. Therefore, aPDT treatment may provide a more favorable healing environment.

This study demonstrated that three important periopathogens are susceptible to aPDT-mediated killing, regardless of whether they are present in planktonic or biofilm form. Furthermore, a clear energy dose-dependence exists with this treatment that should to be taken into account when determining optimal treatment times in clinical application.

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Photodynamic disinfection (PDD) is a nonantibiotic approach to treating drug-resistant bacterial infections. Pseudomonas aeruginosa, an opportunistic pathogen, is problematic because of its propensity to develop antibiotic resistance and its ability to secrete a protective biofilm matrix. This study examined the ability of PDD to eradicate planktonic and biofilm cultures of P. aeruginosa in vitro. Planktonic P. aeruginosa cultures were briefly exposed to a methylene blue-based photosensitizer formulation and subjected to energy doses ranging from 1.7 to 20.6 J cm )2 using a 670 nm nonthermal diode laser. Biofilms were grown for 24 and 48 h and exposed to photosensitizer for 30 s before illumination with 13.2 or 26.4 J of energy. A single exposure of planktonic P. aeruginosa to photosensitizer at >15.5 J cm )2 resulted in 100% eradication (>7 log 10 reduction from control), an effect that could be decreased significantly in the presence of the singlet oxygen quenchers L L-tryptophan and sodium azide. Decreasing the energy dose below this threshold by varying both power density and illumination duration resulted in a dose-dependent decrease in bacterial kill. In addition, 24 h biofilm viability was reduced by 99% with single exposure and 99.9% with double exposure, while 48 h biofilm viability was reduced by >99.999% with both single and double exposures. This study shows that PDD is effective in eradicating planktonic and biofilm cultures of P. aeruginosa, supporting the concept that translation into clinical practice for indications such as otitis externa and wound disinfection is a viable option.

Adult pancreatic ductal cells are believed to be islet precursors. Our aim was to obtain an enriched human ductal cell population in defined culture conditions, and to characterize these cultures for the presence of pancreatic developmental transcription factors. Non-endocrine adult human pancreatic digest was cultured for 4 days in serum-containing and serum-free media. During this time, analysis was done for phenotypic changes, cell death, and expression of islet and islet precursor markers. Culture in serum-supplemented and serum-free media gave similar recoveries of an enriched ductal population after 4 days. Extensive cell death due to apoptosis and necrosis was also observed over this time period. A donor-age dependent expression of pancreatic and duodenal homeobox gene-1 (PDX-1) in ductal cells was seen at 4 days whereby donors <25 yr expressed significantly more than donors >25 yr. Analysis of gene expression by RT-PCR showed the presence of islet developmental transcription factors neuroD, Nkx6.1, and PDX-1, as well as mature islet hormones. While acinar-ductal transdifferentiation of some cells cannot be ruled out, we provide evidence that the predominant mechanism for the derivation of enriched human ductal cultures in our culture conditions is selective acinar cell death. Furthermore, we have shown that ductal cultures from younger donors exhibit greater plasticity through expression of PDX-1, and may be of greater value in attempts to induce islet neogenesis. The presence, however, of insulin and glucagon mRNA indicates that contaminating endocrine cells remain in these cultures and underscores the need to use caution when assessing differentiation potential.