Analysis of Connective Tissue Progenitor Cell Behavior on Polydimethylsiloxane Smooth and Channel Micro-Textures

Growth of human connective tissue progenitor cells (CTPs) was characterized on smooth and microtextured polydimethylsiloxane (PDMS) surfaces. Human bone marrow derived cells were cultured for nine days under conditions promoting osteoblastic differentiation on Smooth PDMS and PDMS Channel microtextures (11 m high, 45 m wide channels, and separated by 5 m wide ridges). Glass tissue culture dish surfaces were used as controls. Cell numbers per colony, cell density within colonies, alignment of cells, area of colonies, and colony shapes were determined as a function of substrate surface topography. An alkaline phosphatase stain was used as a marker for osteoblastic phenotype. CTPs attached, proliferated, and differentiated on all surfaces with cell process lengths of up to 80 m. Cells on the Smooth PDMS and control surfaces spread and proliferated as colonies in proximity to other cells and migrated in random directions creating colonies that covered significantly larger areas (0.96 and 1.05 mm², respectively) than colonies formed on PDMS Channel textures (0.64 mm²). In contrast, cells on PDMS Channel textures spread, proliferated, aligned along the channel axis, and created colonies that were more dense, and with lengths of longest colony axes that were significantly longer (3252 m) than those on the Smooth PDMS (1265 m) and control surfaces (1319 m). Cells on PDMS Channel textures were aligned at an angle of 14.44° relative to the channel axis, and the resulting colonies exhibited a significantly higher aspect ratio (13.72) compared to Smooth PDMS (1.57) and control surfaces (1.51).     

Antibiotic susceptabilities of Pseudomonas aeruginosa isolates derived from patients with cystic fibrosis under aerobic, anaerobic, and biofilm conditions

Recent studies have determined that Pseudomonas aeruginosa can live in a biofilm mode within hypoxic mucus in the airways of patients with cystic fibrosis (CF). P. aeruginosa grown under anaerobic and biofilm conditions may better approximate in vivo growth conditions in the CF airways, and combination antibiotic susceptibility testing of anaerobically and biofilm-grown isolates may be more relevant than traditional susceptibility testing under planktonic aerobic conditions. We tested 16 multidrug-resistant isolates of P. aeruginosa derived from CF patients using multiple combination bactericidal testing to compare the efficacies of double and triple antibiotic combinations against the isolates grown under traditional aerobic planktonic conditions, in planktonic anaerobic conditions, and in biofilm mode. Both anaerobically grown and biofilm-grown bacteria were significantly less susceptible (P < 0.01) to single and combination antibiotics than corresponding aerobic planktonically grown isolates. Furthermore, the antibiotic combinations that were bactericidal under anaerobic conditions were often different from those that were bactericidal against the same organisms grown as biofilms. The most effective combinations under all conditions were colistin (tested at concentrations suitable for nebulization) either alone or in combination with tobramycin (10 microg ml(-1)), followed by meropenem combined with tobramycin or ciprofloxacin. The findings of this study illustrate that antibiotic sensitivities are dependent on culture conditions and highlight the complexities of choosing appropriate combination therapy for multidrug-resistant P. aeruginosa in the CF lung.     

Ex vivo expansion and prophylactic infusion of CMV-pp65 peptide-specific cytotoxic T-lymphocytes following allogeneic hematopoietic stem cell transplantation.

Cytomegalovirus reactivation and infection post-allogeneic hematopoietic stem cell transplant continue to cause morbidity and mortality. Current pharmacologic therapies are limited by side effects. Adoptive transfer of ex vivo generated cytomegalovirus-specific T cells has the potential to restore immunity, prevent cytomegalovirus, and circumvent the need for pharmacologic therapies. We have generated donor-derived cytomegalovirus-specific cytotoxic T cells using dendritic cells pulsed with the HLA-A2 restricted nonapeptide NLVPMVATV (NLV) derived from the cytomegalovirus-pp65 protein. These cytotoxic T cells have been given prophylactically to 9 recipients aged 4 to 65 years on or after day 28 post-allogeneic hematopoietic stem cell transplant. Only 2 of 9 recipients received T cell depletion in vivo or in vitro. There were no immediate adverse reactions to the infusions. During 97-798 days of follow-up, 2 recipients developed cytomegalovirus reactivation; neither developed cytomegalovirus disease or required pharmacotherapy. Three recipients developed acute graft versus host disease after infusion. Two recipients died, 1 from thrombotic thrombocytopenia purpura secondary to cyclosporine, 1 from complications of graft versus host disease. A transient increase in numbers of cytomegalovirus-specific T cells demonstrated by NLV-tetramer binding was seen in 6 recipients. Prophylactic adoptive transfer of NLV-specific T cells is safe and may be effective in preventing cytomegalovirus reactivation.     

Transcription Factor PU.1 Promotes Conventional Dendritic Cell Identity and Function via Induction of Transcriptional Regulator DC-SCRIPT

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Genetic Conservation of Hemagglutinin Gene of H9 Influenza Virus in Chicken Population in Mainland China

The hemagglutinin (HA) genes of 12 H9N2 influenza virus strains isolated from chickens in Mainland China during the period 1995–2002 were genetically analyzed. All the isolates possessed the same amino acid motif -R-S-S-R/G-L- at the cleavage site of HA. Except for the conserved amino acids, as is the case in the other avian influenza viruses, located in the receptor binding site, all of the 12 isolates possessed N at amino acid position 183; A, T, or V at position 190; K at position 137, whereas the representative strains of the other lineage (except Dk/HK/Y280/97-like lineage) virus of H9N2 viruses had H, E, and R at these positions respectively. These could be considered as the partial molecular markers of the H9 viruses isolated from chickens in Mainland China. Phylogenetic analyses showed HA genes of these isolates belonged to that of A/duck/Hong Kong/Y280/97-like virus lineage. No A/quail/Hong Kong/Gl/97-like virus was found in chicken, population since the outbreak of H9N2 influenza in Mainland China in 1992. The available evidence indicates that HA genes of H9 influenza virus circulating in Mainland China during the past years were well conserved.     

Type A behavior and its association with cardiovascular disease prevalence in Blacks and Whites: The Minnesota heart survey

Population-based surveys were conducted in 1985 and 1986 to measure the prevalence of coronary heart disease (CHD) history and risk factors in Black and White adults. Type A behavior was measured by the Jenkins Activity Survey (JAS). JAS scores were associated with age (negatively), education (positively), and sex (men>women) but were largely unrelated to CHD risk factors. Blacks had significantly lower age- and education-adjusted Type A and component scores than Whites, more so formen than women. Univariate analysis indicated that a history of angina and/or heart attack was positively associated with the Type A score in both Blacks and Whites. Following adjustment for known cardiovascular risk factors, Type A score remained positively and significantly associated with CHD prevalence. These findings are consistent with other cross-sectional studies and suggest that Type A behavior, as measured by the JAS, may increase the risk of CHD in both Blacks and Whites. Follow-up of these cohorts may help to clarify the complex relationship of Type A behavior to the risk of CHD.Supported by a National Heart, Lung and Blood Institute Grant (RO1-23727).