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Christopher H Contag

Publications and source records attributed to Christopher H Contag.

At least 37 records · Page 2Linked to original sources
PubMed2006

Systemic effects of orally-administered zinc and tin (IV) metalloporphyrins on heme oxygenase expression in mice.

Some metalloporphyrins (Mps) inhibit heme oxygenase (HO), the rate-limiting enzyme in the production of bilirubin, and are potential compounds for the treatment of neonatal jaundice. We studied the safety and efficacy of Mps following oral administration. Adult HO-1-luc reporter mice were administered 30 micromol/kg body weight of tin mesoporphyrin (SnMP), zinc bis glycol deuteroporphyrin (ZnBG), or zinc protoporphyrin (ZnPP), or vehicle by oral gavage. Bilirubin production was measured as total body carbon monoxide (CO) excretion (VeCO). HO activity was quantitated via CO measurements by gas chromatography. HO-1 protein was determined by Western blot. HO-1 transcription levels were assessed by in vivo bioluminescence imaging. A significant 28% decrease in bilirubin production occurred within 3 h of SnMP treatment and persisted beyond 48 h. Bilirubin production decreased 15% and 9% by 3 h after administration of ZnBG and ZnPP, respectively, but returned to baseline within 48 h. Maximal inhibition of liver, spleen, and intestine HO activity was seen at 3 h with inhibitory effects decreasing in the order: SnMP > or = ZnBG > or = ZnPP. After SnMP treatment, HO-1 transcription increased 5.7-fold after 24 h. Furthermore, liver and spleen HO-1 protein significantly increased 3.7- and 2.0-fold, respectively, after 24 h. HO-1 transcription and protein were not affected in ZnBG- or ZnPP-treated mice. We conclude that the three Mps are absorbed at different rates in the mouse and affect bilirubin production and HO-1 expression in a tissue- and time-dependent manner.

Administration, Oral↗
PubMed2005

Hemin-activated macrophages home to the pancreas and protect from acute pancreatitis via heme oxygenase-1 induction.

Hemin upregulates heme oxygenase-1 (HO-1), a stress-induced enzyme implicated in protection from a variety of injuries while its related isoform HO-2 is constitutively expressed. The role of hemin or HO-1 in the pancreas and their potential modulation of pancreatic injury are unknown. We show that HO-1 is induced in pancreatitis caused by caerulein and more prominently in severe pancreatitis caused by feeding a choline-deficient diet (CDD). Intraperitoneal hemin administration dramatically increases peritoneal and pancreas macrophages that overexpress HO-1 in association with pancreatic induction of the chemoattractants monocyte chemotactic protein-1 and macrophage inflammatory protein-1alpha but not RANTES or macrophage inflammatory protein-2. Hemin administration before CDD feeding protected 8 of 8 mice from lethality while 7 of 16 controls died. Protection is mediated by HO-1-overexpressing macrophages since hemin-primed macrophages home to the pancreas after transfer to naive mice and protect from CDD-induced pancreatitis. Suppression of hemin-primed peritoneal cell HO-1 using HO-1-specific small interfering RNA prior to cell transfer abolishes protection from CDD-induced pancreatitis. Similarly, hemin pretreatment in caerulein-induced pancreatitis reduces serum amylase and lipase, decreases pancreatic trypsin generation, and protects from lung injury. Therefore, hemin-like compounds or hemin-activated macrophages may offer novel therapeutic approaches for preventing acute pancreatitis and its pulmonary complication via upregulation of HO-1.

Acute Disease↗
PubMed2005

The effectiveness of oral tin mesoporphyrin prophylaxis in reducing bilirubin production after an oral heme load in a transgenic mouse model.

BACKGROUND: Neonatal jaundice is commonly encountered and rarely associated with morbidity and mortality. Nonetheless, infants with glucose-6-phosphate dehydrogenase deficiency often have hemolysis (a heme load) caused by an environmental oxidant trigger, thus increasing their risk for serious morbidity. The use of tin mesoporphyrin (SnMP) has been proposed for interdicting the development of severe hyperbilirubinemia in a variety of conditions. OBJECTIVES: We studied the in vivo effects of prophylactic oral SnMP on heme oxygenase (HO) activity and bilirubin production, as indexed by the excretion rate of carbon monoxide (VeCO), following a subsequent oral heme load. METHODS: Adult mice were exposed serially to heme and assessed for in vivo bilirubin production rates, HO-1 transcription and protein, and HO activity. The effect of prophylaxis with a single oral dose of SnMP prior to an oral heme load was assessed by measuring VeCOand tissue HO activities. RESULTS: After serial heme exposures, VeCO, HO-1 transcription and protein, and liver and spleen HO activities increased incrementally. After pretreatment with oral SnMP, bilirubin production decreased in response to an oral heme load. Also, heme-mediated increases in liver, spleen, and intestine HO activities were significantly dampened. CONCLUSIONS: A single oral dose of SnMP results in durable inhibition of bilirubin production and HO activity for at least 24 h in a mouse model of oral heme loading. Further studies are needed to fully elucidate the duration of this protection against hyperbilirubinemia due to a delayed heme load and any long-term consequences of prophylaxis with SnMP on HO-1 transcription and HO-1 protein.

Animals↗
PubMed2005

In vivo imaging of engrafted neural stem cells: its application in evaluating the optimal timing of transplantation for spinal cord injury.

Neural stem/progenitor cells (NSPCs) hold promise in neural tissue replacement therapy after spinal cord injury. However, understanding the survival time of grafted NSPCs and determining the extent of migration away from transplantation sites are essential for optimizing treatment regimens. Here, we used in vivo bioluminescence imaging to noninvasively assess the survival and residence time of transplanted NSPCs at the injury sites in living animals, and we used histologic analyses to assess cell integration and morphology. Third-generation lentiviral vectors enabled efficient transduction and stable expression of both luciferase and a variant of green fluorescent protein in primary cultured NSPCs. Signals from these cells were detectable for up to 10 months or more after transplantation into the injured spinal cords of C57BL/6J mice. Histological and functional data supported the imaging data and suggest that the timing of NSPC transplantation may be a key determinant of the fates and function of integrated cells since cell survival and migration depended on the time of transplantation relative to injury. Optimization of cell therapies can be greatly accelerated and refined by imaging, and the methods in the present study can be widely applied to various research fields of regeneration medicine, including transplantation study.

Animals↗
PubMed2005

Breast-milk shedding of drug-resistant HIV-1 subtype C in women exposed to single-dose nevirapine.

Single-dose nevirapine reduces intrapartum human immunodeficiency virus 1 type (HIV-1) transmission but may also select for nonnucleoside reverse-transcriptase inhibitor (NNRTI) resistance in breast milk (BM) and plasma. Among 32 Zimbabwean women, median 8-week postpartum plasma and BM HIV-1 RNA levels were 4.57 and 2.13 log(10) copies/mL, respectively. BM samples from women with laboratory-diagnosed mastitis (defined as elevated BM Na(+) levels) were 5.4-fold more likely to have HIV-1 RNA levels above the median. BM RT sequences were not obtained for 12 women with BM HIV-1 RNA levels below the lower limit of detection of the assay used. In 20 paired BM and plasma samples, 65% of BM and 50% of plasma RT sequences had NNRTI-resistance mutations, with divergent mutation patterns.

Anti-HIV Agents↗
PubMed2005

Molecular imaging using labeled donor tissues reveals patterns of engraftment, rejection, and survival in transplantation.

Tissue regeneration and transplantation of solid organs involve complex processes that can only be studied in the context of the living organism, and methods of analyzing these processes in vivo are essential for development of effective transplantation and regeneration procedures. We utilized in vivo bioluminescence imaging (BLI) to noninvasively visualize engraftment, survival, and rejection of transplanted tissues from a transgenic donor mouse that constitutively expresses luciferase. Dynamic early events of hematopoietic reconstitution were accessible and engraftment from as few as 200 transplanted whole bone marrow (BM) cells resulted in bioluminescent foci in lethally irradiated, syngeneic recipients. The transplantation of autologous pancreatic Langerhans islets and of allogeneic heart revealed the tempo of transplant degeneration or immune rejection over time. This imaging approach is sensitive and reproducible, permits study of the dynamic range of the entire process of transplantation, and will greatly enhance studies across various disciplines involving transplantation.

Actins↗
PubMed2005

Global analysis of Smad2/3-dependent TGF-beta signaling in living mice reveals prominent tissue-specific responses to injury.

Smad2 and Smad3 (Smad2/3) proteins are key signaling molecules for TGF-beta and some related family members regulating the transcription of several hundred genes. TGF-beta have key roles in development, tissue homeostasis, and the pathogenesis of many human diseases, including cancer, fibrotic disorders, developmental defects, and neurodegeneration. To study the temporal and spatial patterns of Smad2/3-dependent signaling in normal and pathological conditions in the living organism, we engineered transgenic mice with a Smad-responsive luciferase reporter construct (SBE-luc mice). Using bioluminescent imaging, we assessed Smad2/3 signaling activity noninvasively in living mice. At baseline, this activity was highest in brain, intestine, heart, and skin, and correlated with biochemical measurements of reporter activity. Primary astrocytes cultured from SBE-luc mice showed specific activation of the reporter in response to Smad2/3-activating TGF-beta family members. Treatment of mice with the endotoxin LPS resulted in a fast and vigorous, but transient activation of the reporter in the intestine. Although the response was similarly rapid in brain, it remained increased, indicating important but different cellular responses to endotoxin challenge in these organs. Traumatic brain injury with a needle stab resulted in local activation of Smad2/3-dependent genes and a severalfold increase in bioluminescence in living mice. SBE-luc mice can therefore be used to study temporal, tissue-specific activation of Smad2/3-dependent signaling in living mice in normal or pathological conditions as well as for the identification of endogenous or synthetic modulators of this pathway.

Animals↗
PubMed2005

Low levels of Her2/neu expressed by Ewing's family tumor cell lines can redirect cytokine-induced killer cells.

PURPOSE: To identify novel treatments for pediatric solid tumors and/or for malignancies with low-level Her2/neu expression. EXPERIMENTAL DESIGN: Using fluorescence-activated cell sorting and immunohistochemistry, Her2/neu expression was determined on cell lines derived vfrom Ewing's family tumors (EFT) and neuroblastoma. Sensitivity to trastuzumab treatment was investigated using an in vitro proliferation assay. Cytotoxicity against EFT cell lines was done with either freshly isolated or ex vivo activated and expanded T cells (cytokine-induced killer cells, CIK cells), with or without addition of a CD3xHer2/neu bispecific antibody. The effects of either trastuzumab, CIK cells alone, or CD3xHer2/neu bispecific antibody redirected CIK cells was determined using a SCID/hu model of EFTs and serial, noninvasive bioluminescent imaging. RESULTS: EFT cell lines express 5- to 10-fold lower levels of her2/neu than either breast (BT-474) or ovarian (SK-OV-3) cell lines. Treatment of EFT cell lines with trastuzumab did not induce growth inhibition either in vitro or in vivo. In contrast, Her2/neu could be used to redirect CIK cell to mediate cytotoxicity against EFTs both in vitro and in vivo (using two different treatment schemas). CONCLUSIONS: CD3xHer2/neu bispecific antibody and CIK cells may be a suitable approach to treat malignancies with low-level Her2/neu expression not responsive to trastuzumab.

Animals↗
PubMed2005

In vivo analyses of early events in acute graft-versus-host disease reveal sequential infiltration of T-cell subsets.

Graft-versus-host disease (GVHD) is a major obstacle in allogeneic hematopoietic cell transplantation. Given the dynamic changes in immune cell subsets and tissue organization, which occur in GVHD, localization and timing of critical immunological events in vivo may reveal basic pathogenic mechanisms. To this end, we transplanted luciferase-labeled allogeneic splenocytes and monitored tissue distribution by in vivo bioluminescence imaging. High-resolution analyses showed initial proliferation of donor CD4+ T cells followed by CD8+ T cells in secondary lymphoid organs with subsequent homing to the intestines, liver, and skin. Transplantation of purified naive T cells caused GVHD that was initiated in secondary lymphoid organs followed by target organ manifestation in gut, liver, and skin. In contrast, transplanted CD4+ effector memory T (T(EM)) cells did not proliferate in secondary lymphoid organs in vivo and despite their in vitro alloreactivity in mixed leukocyte reaction (MLR) assays did not cause acute GVHD. These findings underline the potential of T-cell subsets with defined trafficking patterns for immune reconstitution without the risk of GVHD.

Animals↗
PubMed2005

Adoptive transfer of mast cells does not enhance the impaired survival of Kit(W)/Kit(W-v) mice in a model of low dose intraperitoneal infection with bioluminescent Salmonella typhimurium.

Mast cells are important effector cells in IgE-associated immune responses, but also can contribute to host defense in certain examples of bacterial infection. We found that genetically mast cell-deficient WBB6F1-Kit(W)/Kit(W-v) mice exhibited more bacterial CFUs per spleen by 6 days after intraperitoneal injection of bioluminescent Salmonella typhimurium, and died more rapidly after infection, than did the congenic WBB6F1-Kit(+/+) wild type mice. Adoptive transfer of bone marrow-derived cultured mast cells of Kit(+/+) origin to the peritoneal cavity of Kit(W)/Kit(W-v) mice resulted in engraftment of mast cells in the peritoneal cavity and mesentery of the recipient mice, and the development of large numbers of mast cells in the spleen. However, such mast cell-engrafted Kit(W)/Kit(W-v) mice appeared sicker after intraperitoneal injection with S. typhimurium than did mast cell-deficient Kit(W)/Kit(W-v) mice, and exhibited numbers of CFUs of bacteria per spleen, and a survival curve, that were not significantly different than those of Kit(W)/Kit(W-v) mice. These results, when taken together with prior studies investigating the roles of mast cells in innate immunity, strongly suggest that whether mast cells can be shown to have a significant role in enhancing survival during bacterial infections may depend critically on the details of the particular experimental systems examined.

Animals↗
PubMed2005

Monitoring age-related susceptibility of young mice to oral Salmonella enterica serovar Typhimurium infection using an in vivo murine model.

Neonates and young children are acutely susceptible to infections by gastrointestinal bacterial pathogens, such as Salmonella enterica serovar Typhimurium (S. typhimurium). To reveal age-related differences in susceptibility to this pathogen, we used in vivo bioluminescence imaging (BLI) to monitor the progression of infection in neonatal (1-wk-old), suckling (2-wk-old), juvenile (4-wk-old), and adult (6-wk-old) BALB/c mice. Mice were orally infected with various doses of a bioluminescent-labeled wild-type or mutant S. typhimurium strain, and progression of infection was monitored by BLI for 2 wks. We found that neonatal and suckling mice were more susceptible to the wild-type strain at inoculum sizes 4 and 2 log(10)'s lower for neonatal and suckling mice, respectively, than those for adult mice. At the lower inocula, newborn mice showed disseminated systemic infection as indicated by the pattern of photon emission assessed by BLI, whereas no bioluminescent signals were detectable in adult mice. In addition, an orgA(-) mutant strain of S. typhimurium with reduced virulence in adult mice produced systemic infection in newborn, suckling, and juvenile mice. Furthermore, as low as 3 log(10) CFU could be detected by BLI in tissue. The present study demonstrates that susceptibility to S. typhimurium infection decreases with age. Also, we established that BLI can be used to monitor the progression of infection in mice. Thus, this model of age-related susceptibility to S. typhimurium using BLI can be used to advance our understanding of the mechanisms involved in newborn susceptibility to infection.

Age Factors↗
PubMed2005

Real-time in vivo imaging of stem cells following transgenesis by transposition.

Previous studies have identified Sleeping Beauty transposons as efficient vectors for nonviral gene delivery in mammalian cells. However, studies demonstrating the usefulness of transposons as gene delivery vehicles into adult stem cells are lacking. Multipotent adult progenitor cells (MAPC) are nonhematopoietic stem cells with the capacity to form most, if not all, cell types of the body and as such hold great therapeutic potential. The whole-body biodistribution and persistence of MAPC are unknown, and such data would help direct clinical applications. We have nucleofected murine MAPC with two plasmid-based Sleeping Beauty transposons encoding the red fluorescent protein (DsRed2) and firefly luciferase. Transgenic euploid MAPC clones maintained their characteristic multilineage differentiation potential in vitro. DsRed2 and luciferase expression allowed for MAPC detection in vivo and in tissue sections. To confirm that transgenesis occurred by transposition into the genome of MAPC, we mapped Sleeping Beauty transposon integration sites in two MAPC clones using splinkerette PCR. This novel dual-reporter imaging approach based on the transgenesis of MAPC with Sleeping Beauty transposons sheds light on the homing patterns of MAPC and paves the way for quantification of MAPC engraftment in real time in vivo.

Animals↗
PubMed2005

Small hairpin RNAs efficiently inhibit hepatitis C IRES-mediated gene expression in human tissue culture cells and a mouse model.

Treatment and prevention of hepatitis C virus (HCV) infections remain a major challenge for controlling this worldwide health problem; existing therapies are only partially effective and no vaccine is currently available. RNA interference offers the potential of a novel therapeutic approach for treating HCV infections. Toward this end, we evaluated small hairpin interfering RNAs (shRNAs) targeting the conserved internal ribosome entry site (IRES) element of the HCV genome for their ability to control gene expression in human cells and animals. We used a reporter gene plasmid in which firefly luciferase (fLuc) expression is dependent on the HCV IRES. Direct delivery of HCV IRES shRNAs efficiently blocked HCV IRES-mediated fLuc expression in transfected human 293FT cells as well as in a mouse model in which nucleic acids were delivered to liver cells by hydrodynamic transfection via the tail vein. These results indicate that shRNAs, delivered as RNA or expressed from viral or nonviral vectors, may be effective agents for the control of HCV and related viruses.

Animals↗
PubMed2005

Ex vivo expanded dendritic cells home to T-cell zones of lymphoid organs and survive in vivo after allogeneic bone marrow transplantation.

Little is known about adoptive transfer of allogeneic ex vivo expanded dendritic cells (eDCs). We investigated the trafficking pattern of eDCs in mice after allogeneic bone marrow transplantation by using bioluminescence imaging. eDCs were expanded from bone marrow precursors in the presence of GM-CSF, interleukin-4, and Flt3L and retrovirally transduced to express luciferase (luc) and green fluorescence protein (gfp). Flow cytometry showed polyclonal DC populations after expansion that consisted of CD11c+CD11b+ and CD11c-CD11b+ cells that co-expressed CD40, CD80, CD86, and MHCII. eDCs were functional in mixed lymphocyte reactions and produced tumor necrosis factor-alpha on phytohemagglutinin stimulation. The eDCs were then injected intravenously into BALB/c recipient mice that had received allogeneic bone marrow transplantation 6 weeks previously. On day 1 after transfer, eDCs were detected by bioluminescence imaging throughout the lungs and spleen. In the later course, signals were observed throughout thymus, lower abdomen, and spleen throughout a period of more than 42 days. Immunofluorescence microscopy confirmed CD11c positivity on the gfp+ donor cells, which localized in T-cell zones of mesenteric lymph nodes, Peyer's patches, spleen, and thymus. These findings are important for adoptive immunotherapies because they indicate that eDCs migrate efficiently in vivo and are capable of surviving long term.

Adjuvants, Immunologic↗
PubMed2004

MYC inactivation uncovers pluripotent differentiation and tumour dormancy in hepatocellular cancer.

Hepatocellular carcinoma is generally refractory to clinical treatment. Here, we report that inactivation of the MYC oncogene is sufficient to induce sustained regression of invasive liver cancers. MYC inactivation resulted en masse in tumour cells differentiating into hepatocytes and biliary cells forming bile duct structures, and this was associated with rapid loss of expression of the tumour marker alpha-fetoprotein, the increase in expression of liver cell markers cytokeratin 8 and carcinoembryonic antigen, and in some cells the liver stem cell marker cytokeratin 19. Using in vivo bioluminescence imaging we found that many of these tumour cells remained dormant as long as MYC remain inactivated; however, MYC reactivation immediately restored their neoplastic features. Using array comparative genomic hybridization we confirmed that these dormant liver cells and the restored tumour retained the identical molecular signature and hence were clonally derived from the tumour cells. Our results show how oncogene inactivation may reverse tumorigenesis in the most clinically difficult cancers. Oncogene inactivation uncovers the pluripotent capacity of tumours to differentiate into normal cellular lineages and tissue structures, while retaining their latent potential to become cancerous, and hence existing in a state of tumour dormancy.

Animals↗
PubMed2004

Adipose-derived adult stromal cells heal critical-size mouse calvarial defects.

In adults and children over two years of age, large cranial defects do not reossify successfully, posing a substantial biomedical burden. The osteogenic potential of bone marrow stromal (BMS) cells has been documented. This study investigates the in vivo osteogenic capability of adipose-derived adult stromal (ADAS) cells, BMS cells, calvarial-derived osteoblasts and dura mater cells to heal critical-size mouse calvarial defects. Implanted, apatite-coated, PLGA scaffolds seeded with ADAS or BMS cells produced significant intramembranous bone formation by 2 weeks and areas of complete bony bridging by 12 weeks as shown by X-ray analysis, histology and live micromolecular imaging. The contribution of implanted cells to new bone formation was 84-99% by chromosomal detection. These data show that ADAS cells heal critical-size skeletal defects without genetic manipulation or the addition of exogenous growth factors.

Adipose Tissue↗
PubMed2004

The E47 transcription factor negatively regulates CD5 expression during thymocyte development.

The expression of CD5 increases progressively as thymocytes mature. We have shown that CD5 expression is controlled by a tissue-specific regulatory promoter located upstream of the CD5 translation start sites. Deletion of this regulatory promoter, which contains three potential transcription factor binding sites (CCAAT, kappa E2, and ets) reduces the promoter activity to basal level. Of these sites, only ets proved essential for CD5 expression in T cell lines. Here, we introduce a role for the E47 transcription factor and the CD5 promoter kappa E2 site in regulating CD5 expression during thymocyte development. Using T cell lines, we show that (i) mutation of the kappa E2 site in the CD5 regulatory promoter results in a significant elevation of CD5 promoter activity; (ii) the E47 transcription factor binds to the kappa E2 site; and (iii) overexpression of E47 inhibits CD5 expression. We then show, in high-dimensional fluorescence-activated cell sorting studies with primary thymocytes at successive developmental stages, that (i) intracellular E47 levels decrease as surface CD5 expression increases; (ii) E47 expression is down-regulated and CD5 expression is correspondingly up-regulated in DN3 thymocytes in RAG-2-deficient mice injected with anti-CD3 to mimic pre-T cell receptor stimulation; and (iii) E47 expression is down-regulated and CD5 expression is up-regulated when double positive thymocytes are stimulated in vitro with anti-CD3. Based on these data, we propose that E47 negatively regulates CD5 expression by interacting with the kappa E2 site in the CD5 regulatory promoter and that decreases in E47 in response to developmental signals are critical to the progressive increase in CD5 expression as thymocytes mature.

Animals↗