Housing characteristics and indoor air quality in households of Alaska Native children with chronic lung conditions.

Alaska Native children experience high rates of respiratory infections and conditions. Household crowding, indoor smoke, lack of piped water, and poverty have been associated with respiratory infections. We describe the baseline household characteristics of children with severe or chronic lung disease participating in a 2012-2015 indoor air study. We monitored indoor PM2.5, CO2 , relative humidity %, temperature, and VOCs and interviewed caregivers about children's respiratory symptoms. We evaluated the association between reported children's respiratory symptoms and indoor air quality indicators using multiple logistic regression analysis. Compared with general US households, study households were more likely overcrowded 73% (62%-82%) vs 3.2% (3.1%-3.3%); had higher woodstove use as primary heat source 16% (9%-25%) vs 2.1% (2.0%-2.2%); and higher proportion of children in a household with a smoker 49% (38%-60%) vs 26.2% (25.5%-26.8%). Median PM2.5 was 33 µg/m3 . Median CO2 was 1401 ppm. VOCs were detectable in all homes. VOCs, smoker, primary wood heat, and PM2.5>25 µg/m3 were associated with higher risk for cough between colds; VOCs were associated with higher risk for wheeze between colds and asthma diagnosis. High indoor air pollutant levels were associated with respiratory symptoms in household children, likely related to overcrowding, poor ventilation, woodstove use, and tobacco smoke.

The Strong Family Program: an innovative model to engage Aboriginal and Torres Strait Islander youth and Elders with reproductive and sexual health community education.

Issue addressed Aboriginal youth in Australia often experience high rates of intimate partner violence (family violence) and poorer reproductive and sexual health than their non-Aboriginal counterparts. To address some of the disparities, the Strong Family Program was developed to deliver reproductive and sexual health education to Aboriginal communities in New South Wales. Methods Development of the program was based on an extensive consultation process with Aboriginal communities. It was implemented in three communities, with two groups from each hosting Aboriginal youth and Elders in a yarning circle within the culturally respectful frameworks of 'men and boys'' and 'women and girls'' business. An evaluation was conducted to measure reproductive and sexual health knowledge and attitude changes upon program completion, using pre- and post-program surveys and yarning (focus group discussions). Results Program participants comprised 48 females and 28 males. Overall, mean knowledge and attitude scores improved upon completion of the program (from 77% to 82% and from 4.15 to 4.32 out of 5, respectively). Among participants aged 20 years and under (the youngest participant was 13 years), there was an increase in knowledge (P=0.034); among participants aged over 20 years (the oldest participant was 78 years), there was an increase in positive attitudes (P=0.001). Participants perceived the information provided to be useful and relevant, with many reporting improved knowledge and attitudes around rights and respectful relationships. Conclusions Reproductive and sexual health education in Aboriginal communities should be based on community consultations and carried out within a culturally appropriate framework to promote greater success. Continued implementation of the Strong Family Program will promote increased understanding of respectful relationships and improved health outcomes for Aboriginal young people. So what? The Strong Family Program was based on an extensive consultative process that ensured leadership and involvement from Aboriginal communities, with program content and delivery based on Aboriginal pedagogy and reflecting Aboriginal cultural values. Reproductive and sexual health promotion in Aboriginal communities should be based on community consultations and carried out within a culturally appropriate framework to promote greatest success.

Technical Report: Evaluation of peripheral dose for flattening filter free photon beams.

PURPOSE: To develop a comprehensive peripheral dose (PD) dataset for the two unflattened beams of nominal energy 6 and 10 MV for use in clinical care. METHODS: Measurements were made in a 40 × 120 × 20 cm(3) (width × length × depth) stack of solid water using an ionization chamber at varying depths (dmax, 5, and 10 cm), field sizes (3 × 3 to 30 × 30 cm(2)), and distances from the field edge (5-40 cm). The effects of the multileaf collimator (MLC) and collimator rotation were also evaluated for a 10 × 10 cm(2) field. Using the same phantom geometry, the accuracy of the analytic anisotropic algorithm (AAA) and Acuros dose calculation algorithm was assessed and compared to the measured values. RESULTS: The PDs for both the 6 flattening filter free (FFF) and 10 FFF photon beams were found to decrease with increasing distance from the radiation field edge and the decreasing field size. The measured PD was observed to be higher for the 6 FFF than for the 10 FFF for all field sizes and depths. The impact of collimator rotation was not found to be clinically significant when used in conjunction with MLCs. AAA and Acuros algorithms both underestimated the PD with average errors of -13.6% and -7.8%, respectively, for all field sizes and depths at distances of 5 and 10 cm from the field edge, but the average error was found to increase to nearly -69% at greater distances. CONCLUSIONS: Given the known inaccuracies of peripheral dose calculations, this comprehensive dataset can be used to estimate the out-of-field dose to regions of interest such as organs at risk, electronic implantable devices, and a fetus. While the impact of collimator rotation was not found to significantly decrease PD when used in conjunction with MLCs, results are expected to be machine model and beam energy dependent. It is not recommended to use a treatment planning system to estimate PD due to the underestimation of the out-of-field dose and the inability to calculate dose at extended distances due to the limits of the dose calculation matrix.

Impact of providing in-home water service on the rates of infectious diseases: results from four communities in Western Alaska.

Approximately 20% of rural Alaskan homes lack in-home piped water; residents haul water to their homes. The limited quantity of water impacts the ability to meet basic hygiene needs. We assessed rates of infections impacted by water quality (waterborne, e.g. gastrointestinal infections) and quantity (water-washed, e.g. skin and respiratory infections) in communities transitioning to in-home piped water. Residents of four communities consented to a review of medical records 3 years before and after their community received piped water. We selected health encounters with ICD-9CM codes for respiratory, skin and gastrointestinal infections. We calculated annual illness episodes for each infection category after adjusting for age. We obtained 5,477 person-years of observation from 1032 individuals. There were 9,840 illness episodes with at least one ICD-9CM code of interest; 8,155 (83%) respiratory, 1,666 (17%) skin, 241 (2%) gastrointestinal. Water use increased from an average 1.5 gallons/capita/day (g/c/d) to 25.7 g/c/d. There were significant (P-value < 0.05) declines in respiratory (16, 95% confidence interval (CI): 11-21%), skin (20, 95%CI: 10-30%), and gastrointestinal infections (38, 95%CI: 13-55%). We demonstrated significant declines in respiratory, skin and gastrointestinal infections among individuals who received in-home piped water. This study reinforces the importance of adequate quantities of water for health.

Culture expanded primary chondrocytes have potent immunomodulatory properties and do not induce an allogeneic immune response.

OBJECTIVE: Allogeneic cell therapies, such as mesenchymal stromal cells (MSC), which have potent regenerative and anti-inflammatory potential are being investigated as a therapy for osteoarthritis (OA) and cartilage injury. Here we describe another potential source of regenerative and anti-inflammatory allogeneic cells, culture expanded primary chondrocytes (CEPC). In direct comparison to allogeneic MSC, we extensively assess the immunological interactions of CEPC in an allogeneic setting. METHODS: Chondrocytes were isolated from rat articular cartilage and cultured in normoxic or hypoxic conditions. In vitro co-culture assays with allogeneic lymphocytes and macrophages were used to assess the immunomodulatory capacities of the chondrocytes, followed by immune response analysis by flow cytometry, ELISA and qPCR. RESULTS: CEPC showed reduced induction of proliferation, activation and cytotoxic granzyme B expression in allogeneic T cells. Importantly, exposure to pro-inflammatory cytokines did not increase CEPC immunogenicity despite increases in MHC-I. Furthermore, CEPC had a potent ability to suppress allogeneic T cell proliferation, which was dependent on nitric oxide production. This suppression was contact independent in hypoxia cultured CEPC. Finally, chondrocytes were shown to have the capacity to modulate pro-inflammatory macrophage activity by reducing MHC-II expression and TNF-α secretion. CONCLUSION: These data indicate the potential use of allogeneic chondrocytes in OA and cartilage defects. The lack of evident immunogenicity, despite exposure to a pro-inflammatory environment, coupled with the immunomodulatory ability indicates that these cells have the potential to evade the host immune system and suppress inflammation, thus potentially facilitating the resolution of OA induced inflammation and cartilage regeneration.

Mesenchymal stem cell therapy promotes corneal allograft survival in rats by local and systemic immunomodulation.

Mesenchymal stem cells (MSCs) are being investigated extensively due to their ability to dampen immune responses. Here, we tested the ability of MSCs from three distinct sources to prolong rat corneal allograft survival. A fully allogeneic rat cornea transplant model (DA to LEW) was used. Recipient rats received 1 × 10(6) MSCs (syn [LEW], allo [DA] or third-party [Wistar Furth]) intravenously 7 days before transplantation and again on the day of transplantation (day 0). A high percentage of untreated and syn-MSC treated allografts were rejected (80% and 100%, respectively). Preactivation of syn-MSCs with interferon gamma also failed to prolong allograft survival. Conversely, corneal allograft survival was significantly prolonged in allo-MSC treated (90%) and third-party MSC treated (80%) allograft recipients. Flow cytometric analysis revealed less infiltrating natural killer T cells in corneas of both allo- and third-party MSC treated animals, coupled with a higher proportion of splenic CD4+Foxp3+ regulatory T cells, compared to controls. In the case of allo- and third-party MSCs, results from a delayed-type hypersensitivity assay clearly showed that hypo-responsiveness was specific for corneal donor-associated allo-antigens. Thus, allo- and third-party MSC treatment prolongs corneal allograft survival by suppressing peripheral immune responses and promoting an intragraft immunoregulatory milieu.

Role of lentivirus-mediated overexpression of programmed death-ligand 1 on corneal allograft survival.

To investigate the role of lentivirus-mediated overexpression of programmed death-ligand 1 (PD-L1) on rat corneal allograft survival. A fully allogeneic rat cornea transplant model was used for in vivo studies. Lentiviral (LV) vectors are efficient tools for ex vivo genetic modification of cultured corneas. LV vector encoding for PD-L1 (LV.PD-L1) and LV vector encoding for eGFP (LV.eGFP, as control) were constructed and tested. PD-L1 or eGFP expression was increased on corneal cells upon LV.PD-L1 and LV.eGFP transduction, respectively. Both allogeneic controls and allogeneic LV.eGFP transduced corneas were uniformly rejected (MST: 13.8 ± 1.7 days and 12.3 ± 1.9 days, respectively). In contrast, allogeneic LV.PD-L1 transduced corneas showed a high percentage (83%) of graft survival (MST > 30 days, n = 5, 15 days, n = 1). Graft opacity of PD-L1 transduced corneas was present but was significantly reduced compared to control or eGFP expressing corneas. Flow cytometric analysis revealed that percentages of CD3(+) CD8(+) CD161(+) and CD3(+) CD8(+) CD161(-) lymphocytes were decreased in animals receiving LV.PD-L1 transduced corneas compared to animals grafted with LV.eGFP transduced corneas. Moreover, reduced expression of proinflammatory cytokines (IFN-γ and IL-6) in PD-L1 transduced corneas compared to allogeneic controls was also observed. Local PD-L1 gene transfer in cultured corneas is a promising approach for the prolongation of corneal allograft survival and attenuation of graft rejection.

Cytokine-induced beta-cell apoptosis is NO-dependent, mitochondria-mediated and inhibited by BCL-XL.

Pro-inflammatory cytokines are implicated as the main mediators of beta-cell death during type 1 diabetes but the exact mechanisms remain unknown. This study examined the effects of interleukin-1beta (IL-1beta), interferon-gamma (IFNgamma) and tumour necrosis factor alpha (TNFalpha) on a rat insulinoma cell line (RIN-r) in order to identify the core mechanism of cytokine-induced beta-cell death. Treatment of cells with a combination of IL-1beta and IFNgamma (IL-1beta/IFNgamma)induced apoptotic cell death. TNFalpha neither induced beta-cell death nor did it potentiate the effects of IL-1beta, IFNgamma or IL-1beta/IFNgamma . The cytotoxic effect of IL-1beta/IFNgamma was associated with the expression of inducible nitric oxide synthase (iNOS) and production of nitric oxide. Adenoviral-mediated expression of iNOS (AdiNOS) alone was sufficient to induce caspase activity and apoptosis. The broad range caspase inhibitor, Boc-D-fmk, blocked IL-1beta/IFNgamma -induced caspase activity, but not nitric oxide production nor cell death. However, pre-treatment with L-NIO, a NOS inhibitor, prevented nitric oxide production, caspase activity and reduced apoptosis. IL-1beta/IFNgamma -induced apoptosis was accompanied by loss of mitochondrial membrane potential, release of cytochrome c and cleavage of pro-caspase-9, -7 and -3. Transduction of cells with Ad-Bcl-X(L) blocked both iNOS and cytokine-mediated mitochondrial changes and subsequent apoptosis, downstream of nitric oxide. We conclude that cytokine-induced nitric oxide production is both essential and sufficient for caspase activation and beta-cell death, and have identified Bcl-X(L) as a potential target to combat beta-cell apoptosis.

Effects of local and systemic viral interleukin-10 gene transfer on corneal allograft survival.

In this study, we explored the immunomodulatory effects of viral interleukin (IL) IL-10 after ex vivo and in vivo gene transfer in experimental corneal transplantation. Wistar-Furth rats were used as donors and major histocompatibility complex class I/II-disparate Lewis rats served as recipients. For ex vivo gene therapy donor corneas were either transfected with liposome/vIL-10 plasmid DNA mixtures or transduced with a vIL-10 expressing adenovirus vector (AdvIL-10). For in vivo studies, recipients were treated with AdvIL-10 intraperitoneally 1 day before transplantation. Graft survival was analysed using the Kaplan-Meier survival method. To monitor the efficacy of the therapy messenger RNA (mRNA) cytokine expression profiles in grafts and draining lymph nodes were analysed by quantitative real-time reverse transcription-polymerase chain reaction. Moreover, anti-adenovirus immunity was also investigated. Neither ex vivo liposome-mediated vIL-10 gene transfer nor ex vivo AdvIL-10 gene transfer led to prolonged corneal allograft survival. In contrast, corneal allograft survival was significantly prolonged in animals receiving systemic AdvIL-10 gene transfer. Moreover, only systemic vIL-10 gene therapy modulated the cytokine mRNA expression profile in draining lymph nodes. Interestingly, systemic AdvIL-10 gene transfer could not inhibit the generation of anti-adenovirus antibodies. Our data indicate systemic expression of the vIL-10 gene is required to modulate the cytokine expression profile in the draining lymph nodes, which might be a pre-requisite for the success of cytokine gene therapy.

Gene therapy for the prevention of ischemia/reperfusion injury in organ transplantation.

Introduction of gene therapy into molecular medicine has been gaining increasing interest. Although treatment of various diseases e.g. monogenetic defects or cancer by using gene transfer technologies has been extensively probed, the clinical success has been limited. However, recent experimental data suggest that gene therapy may represent an attractive and powerful approach in preventing ischemia/reperfusion injury as well as organ rejection in transplant recipients. Easy and selective access to the donor organ facilitates the reduction of potentially harmful systemic side effects of gene therapy vectors. By introducing anti-apoptotic or cytoprotective genes, these studies focused on the protection of the transplant from the apoptotic cell death. In addition, down-regulation of adhesion molecules and/or blockade of gene expression in the graft itself also ameliorated ischemia/reperfusion injury. This review summarizes the current progress on gene therapy application in combating ischemia-reperfusion injury in organ transplantation. Although the use of viral vectors is emphasized, non-viral gene transfer techniques are also discussed. Future development of novel, low-immunogenic vectors should further contribute to the minimization of ischemia/reperfusion injury, and thus to the overall success of organ transplantation.

Intrabody-mediated phenotypic knockout of major histocompatibility complex class I expression in human and monkey cell lines and in primary human keratinocytes.

Cultured keratinocyte allografts from unrelated donors can be readily grown as sheets in large-scale cell culture and have been used as an immediate skin cover for severely burned patients. Despite the absence of passenger leukocytes and the unlimited amount of material that can be obtained for permanent skin coverage, the allografts are susceptible to rejection. Since MHC class I (MHCI) antigens serve as targets for allograft rejection, we investigated whether 'phenotypic knockout' of human MHCI could be achieved through expression of an ER-directed anti-human MHCI single-chain intrabody (sFvhMHCI) that is directed against a monomorphic, conformational epitope, expressed across species lines, on the MHCI heavy chain. Co-immunoprecipitation of both MHCI heavy chain and beta2-microglobulin occurred in transfected monkey COS-1 cells, while Jurkat T cells stably expressing the ER-directed sFvhMHCI intrabody showed that complete phenotypic knockout of MHCI cell surface expression could be achieved. Infection of several human cell lines of divergent tissue sources and different HLA haplotypes resulted in marked down-regulation of MHCI expression, even under conditions where inflammatory cytokines (eg gamma-IFN) which up-regulate MHCI expression were used. Finally, when adenovirus encoding the anti-human MHCI intrabody was used to transduce primary human keratinocytes, a marked reduction of surface MHCI expression was observed. These in vitro studies set the groundwork for in vivo studies to determine if intrabody-mediated knockout of MHCI can impair alloantigen expression and prolong the survival of keratinocyte allografts.

Fibroblast-mediated delivery of growth factor complementary DNA into mouse joints induces chondrogenesis but avoids the disadvantages of direct viral gene transfer.

OBJECTIVE: To assess the advantages and disadvantages of a direct adenoviral and a cell-mediated approach to the induction of cartilage formation in joints by transfer of growth factor genes. METHODS: Adenoviral vectors carrying insulin-like growth factor 1 (IGF-1) or bone morphogenetic protein 2 (BMP-2) complementary DNA were constructed and applied to primary human and murine chondrocytes or fibroblasts. Transgene expression was quantified by enzyme-linked immunosorbent assay. Direct injection of these vectors or AdLacZ, a reporter gene vector, into mouse knee joints was compared with the transplantation of syngeneic fibroblasts (infected ex vivo with the same vectors) with respect to virus spread, immune response, and cartilage formation by use of histologic, immunohistochemical, and molecular analyses. RESULTS: AdIGF-1 and AdBMP-2 efficiently infected all cell types tested. Human cells secreted biologically relevant levels of protein over a period of at least 28 days. Direct transfer of AdLacZ into mouse knee joints resulted in positively stained synovial tissues, whereas AdLacZ-infected fibroblasts settled on the surface of the synovial membranes. Inadvertent spread of vector DNA into the liver, lung, and spleen was identified by nested polymerase chain reaction in all mice that had received the vector directly; this rarely occurred following fibroblast-mediated gene transfer. Direct injection of AdBMP-2 induced the synthesis of new cartilage in periarticular mesenchyme, accompanied by extensive osteophyte formation. When AdBMP-2 was administered by injecting ex vivo-infected fibroblasts, cartilage formation was observed only in regions near the injected cells. AdIGF-1 treatment did not lead to morphologic changes. Importantly, fibroblast-mediated gene transfer avoided the strong immune response to adenovirus that was elicited following direct application of the vector. CONCLUSION: Our results indicate that cell-mediated gene transfer provides sufficient BMP-2 levels in the joint to induce cartilage formation while avoiding inadvertent vector spread and immune reactions.

Cytokine mRNA levels and lymphocyte infiltration in pancreatic tissue during experimental chronic pancreatitis induced by dibutyltin dichloride.

There is little information available regarding the role of inflammatory cells in the pathogenesis of chronic pancreatitis. Therefore, we analyzed the local cytokine profile and infiltrating lymphocytes in a rat model of chronic pancreatitis. Experimental pancreatitis was induced by a single intravenous application of dibultyltin dichloride (DBTC). During a time course of two months we observed the mRNA expression of cytokines using competitive RT-PCR. Lymphocytes were characterized by immunohistochemistry, FACS analysis, and the lymphocyte proliferation test. IL-1beta, IL-6, IL-5, and IL-10 were immediately up-regulated in the acute phase of disease, while lymphocyte-restricted expression of IL-2, IL-2R, and IFN-y was only found in the chronic course. Among the infiltrating lymphocytes, CD4+ cells dominated, but during the chronic process there was an increase of CD8+ cells, resulting in a reduced CD4/CD8 ratio. Mitogen-induced activation of isolated mesenteric lymph node cells increased during the chronic inflammation. Our results suggest that in experimental pancreatitis acute inflammatory reactions are followed by a T-lymphocyte-mediated process.

Corneal allograft rejection: current understanding. I. Immunobiology and basic mechanisms.

Allograft rejection remains the single largest impediment to success in corneal transplantation. This article briefly reviews our current understanding of some fundamental aspects of corneal immunology and the pathogenetic mechanisms underlying corneal graft rejection. As knowledge increases, it is hoped that a better understanding of the immunobiology may result in improved preventive and therapeutic measures.

High frequency properties of passive materials for ultrasonic transducers.

The acoustic properties of passive materials for ultrasonic transducers have been measured at room temperature in the frequency range from 25 to 65 MHz using ultrasonic spectroscopy. These materials include alumina/EPO-TEK 301 composites and tungsten/EPO-TEK 301 composites. Experimental results showed that the acoustic impedance of the composites monotonically increased with the volume fraction of the particle filler, which is in agreement with the Denavey model. The attenuation, however, peaked between 7 and 9% volume fraction of particle filler. For comparison, several other passive materials were also fabricated and measured. The results suggest that materials that possess a higher attenuation also appear to have a larger velocity dispersion.

Pre-focused lead titanate > 25 MHz single-element transducers from hollow spheres.

Miniature lead titanate (PT) hollow spheres with diameters in the 1 to 10 mm range and wall thicknesses of 20 to 120 microns have been fabricated. Shell sections were used as components of pre-focused transducers. Spheres are produced using a new sacrificial core technique that produces hundreds of spheres with a more uniform wall thickness than those produced by earlier methods. Shells produced from these spheres were found to have a wall thickness variation of about 10%. Despite this variation, bulk properties were estimated from capacitance and impedance data. Shells tested in this work had dielectric constants (1 kHz) near 280 with loss factor of < 2% and d33 values of 68 pC/N. Thickness coupling coefficients averaged 0.51 with mechanical quality factors of < 15. A transducer fabricated from these sections of spheres had a round-trip insertion loss of -20.1 dB at the center frequency of 39.8 MHz and a 6 dB bandwidth of 33%.

Migratory activity and functional changes of green fluorescent effector cells before and during experimental autoimmune encephalomyelitis.

Homing behavior and function of autoimmune CD4+ T cells in vivo was analyzed before and during EAE, using MBP-specific T cells retrovirally engineered to express the gene of green fluorescent protein. The cells migrate from parathymic lymph nodes to blood and to the spleen. Preceding disease onset, large numbers of effector cells invade the CNS, with only negligible numbers left in the periphery. In early EAE, most (>90%) infiltrating CD4+ cells were effector cells. Migratory effector cells downregulate activation markers (CD25, OX-40) but upregulate several chemokine receptors and adsorb MHC class II on their membranes. Within the CNS, the effector cells are reactivated, with upregulated proinflammatory cytokines and downmodulated T cell receptor-associated structures, presumably reflecting autoantigen recognition in situ.