Biomedical subjects

Ronald G Crystal

Publications and source records attributed to Ronald G Crystal.

94 records · Page 6Linked to original sources

PubMed2002

Apoptosis induced by Pseudomonas aeruginosa in antigen presenting cells is diminished by genetic modification with CD40 ligand.

Persistent colonization with Pseudomonas aeruginosa (PA) is a hallmark of the lung disease associated with cystic fibrosis (CF). Based on the concept that PA is not cleared from the lung by the host response in individuals with CF, we analyzed the capacity of PA to induce cell death in human alveolar macrophages (AM) and murine dendritic cells (DC), antigen presenting cells that play a central role in the initiation of pulmonary host defenses against pathogens, and evaluated if genetic modification can lead to protection against PA induced cell death. AM and DC were susceptible to cell death induced by the laboratory PA isolates PAO1, PAK and PA103, as well as a mucoid derivative of PAO1 and PA isolates derived from sputum of individuals with CF. Apoptosis, analyzed by TUNEL assay, was detectable in AM and DC as early as 3 h after infection with PA. In contrast, the same strains and doses of PA had little effect on the lung epithelial cell line A549 and primary cultures of human bronchial epithelial cells in vitro. Pretreatment of DC with the caspase inhibitors VAD-fmk and YVAD-cmk reduced PA induced cell death (p < 0.05). Finally, genetic modification of DC to express CD40L using an adenovirus vector decreased the susceptibility of DC to cell death induced by PAO1 compared with DC infected with a control Ad vector (p < 0.01). The data demonstrate that DC and AM are susceptible to apoptosis induced by PA and that this response can be partially reversed by genetic modification with CD40L, a CD4+ T cell molecule that plays a central role in activating antigen presenting cells. These observations suggest a potential mechanism contributing to the persistence of PA in CF and suggest that genetic manipulation of antigen presenting cells with anti-apoptotic genes may be able to strengthen host defenses in CF.

Adenoviruses, Human↗

PubMed2002

Adenovirus vector-mediated overexpression of a truncated form of the p65 nuclear factor kappa B cDNA in dendritic cells enhances their function resulting in immune-mediated suppression of preexisting murine tumors.

PURPOSE: The purpose of this study was to evaluate the effect of the Rel homology domain (RHD) of the transcription factor, nuclear factor kappa B (NF kappa B), on proinflammatory gene expression in bone marrow-derived dendritic cells (BMDCs). EXPERIMENTAL DESIGN: We used an adenovirus vector encoding only the RHD of the NF kappa B (p65 family member) cDNA (AdRHD) to transduce murine BMDCs ex vivo. Endpoints measured included BMDC expression of activation markers, cytokine secretion, peptide antigen presentation, and the ability of these transduced cells to induce antitumor immunity in vivo. RESULTS: AdRHD-transduced BMDCs secreted higher levels of the cytokines interleukin (IL) 1 beta, IL-6, and IL-12 (p40) compared with sham-transduced BMDCs or those transduced with an empty vector. AdRHD induced heightened surface expression of the activation markers CD40, B7.1, B7.2, and MHC class II on BMDCs, and these cells were able to present a peptide antigen to a T-lymphocyte hybridoma more efficiently than controls in vitro. Growth of syngeneic, established tumors (CT26 and B16.F10) was inhibited, and survival was prolonged in the mice that received intratumoral AdRHD-modified BMDCs compared with controls. Splenocytes from CT26 tumor-bearing animals that received intratumoral AdRHD-modified BMDCs were able to lyse CT26 target cells more efficiently than controls. Similar experiments using host mice harboring targeted mutations in CD4 and CD8, as well as BMDCs from mice lacking MHC class I, MHC class II, or IL-12 revealed that this tumor immunity was dependent on the presence of CD4+ and CD8+ cells in the tumor-bearing host, as well as MHC class I, MHC class II, and IL-12 expression by the administered BMDCs. Furthermore, induction of IL-12 (p40) expression by AdRHD was completely abrogated in BMDCs lacking the c-Rel NF kappa B family member. CONCLUSIONS: We made the following conclusions: (a) gene transfer-mediated overexpression of the RHD of NF kappa B activates BMDCs; (b) AdRHD-transduced BMDCs induce antitumor immunity when administered intratumorally, an effect mediated by both the CD4+ T cell/MHC class II and the CD8+ T cell/MHC class I pathways, as well as IL-12; and (c) IL-12 (p40) up-regulation by the RHD transgene in BMDCs is dependent on the presence of the c-Rel NF kappa B family member.

Adenoviridae↗

PubMedDate not supplied

Intratumoral expression of macrophage-derived chemokine induces CD4+ T cell-independent antitumor immunity in mice.

Macrophage-derived chemokine is chemotactic for a variety of leukocytes, and has been shown to be involved in T 2-mediated cellular immunity. To evaluate the role of this chemokine in tumor immunity in vivo, an adenovirus vector encoding the human macrophage-derived chemokine cDNA (AdMDC) was administered to established murine tumors. Gene transfer with AdMDC significantly inhibited tumor growth and prolonged animal survival. AdMDC was not directly cytotoxic to tumor cells, but splenocytes from animals that received intratumoral AdMDC were able to lyse syngeneic tumor cells, and purified splenic CD8 cells secreted interferon-gamma in a tumor-specific manner. The antitumor activity of AdMDC was lost in mice lacking CD8 T lymphocytes, but surprisingly, it was preserved in animals lacking CD4 cells, as was the systemic cytotoxic T lymphocyte response. Systemic NK cells did not play a role in the antitumor immune response induced by AdMDC. Experiments using knockout mice demonstrated that host expression of MHC Class I, but not Class II, IL-4, or IL-12, was necessary for AdMDC to exert its antitumor effect, and immunohistochemistry demonstrated infiltrates of CD8 and CD86 cells, but not CD4 cells in treated tumors. These studies highlight a new function for macrophage-derived chemokine by demonstrating that it possesses in vivo antitumor activity with CD8 T cells as the effector cells, and interestingly, that the CD4 cell/MHC II pathway of CD8 cell activation is not required for the antitumor effects of this chemokine.(H)

Adenocarcinoma↗

PubMedDate not supplied

Sampling-dependent up-regulation of gene expression in sequential samples of human airway epithelial cells.

As part of a study of in vivo gene expression levels in the human airway epithelium in response to chronic cigarette smoking, we have identified a number of genes whose expression levels are altered in a time-dependent fashion resulting from the procedure used to sample epithelial cells. Fiberoptic bronchoscopy and airway epithelium brushing were used to obtain independent samples from a single individual, 1st from the right lung, followed by sampling of the left lung. We observed that a specific subset of early response genes encoding proteins involved in transcription, signal transduction, cell cycle/growth, and apoptosis were significantly up-regulated in the left lung samples (the 2nd region to be sampled) compared with the right lung samples (the 1st region to be sampled). This response was due to the temporal nature of the sampling procedure and not to inherent gene expression differences between airway epithelium of the right and left lungs. When the order of sampling was reversed, with the left airway epithelium sampled 1st, the same subset of genes were up-regulated in the samples obtained from the right airway epithelium. The time-dependent up-regulation of these genes was likely in response to the stress of the procedure and/or the anesthesia used. Sampling-dependent uncertainty of gene expression is likely a general phenomenon relevant to the procedures used for obtaining biological samples, particularly in humans where the sampling procedures are dependent on ensuring comfort and safety.

Apoptosis↗