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Biomedical subjects

P P Smith

Publications and source records attributed to P P Smith.

At least 19 recordsLinked to original sources
PubMed2001

Reactivation of latent human cytomegalovirus in CD14(+) monocytes is differentiation dependent.

We have previously demonstrated reactivation of latent human cytomegalovirus (HCMV) in myeloid lineage cells obtained from healthy donors. Virus was obtained from allogenically stimulated monocyte-derived macrophages (Allo-MDM), but not from macrophages differentiated by mitogenic stimulation (ConA-MDM). In the present study, the cellular and cytokine components essential for HCMV replication and reactivation were examined in Allo-MDM. The importance of both CD4(+) and CD8(+) T cells in the generation of HCMV-permissive Allo-MDM was demonstrated by negative selection or blocking experiments using antibodies directed against both HLA class I and HLA class II molecules. Interestingly, contact of monocytes with CD4 or CD8 T cells was not essential for reactivation of HCMV, since virus was observed in macrophages derived from CD14(+) monocytes stimulated by supernatants produced by allogeneic stimulation of peripheral blood mononuclear cells. Examination of the cytokines produced in Allo-MDM and ConA-MDM cultures indicated a significant difference in the kinetics of production and quantity of these factors. Further examination of the cytokines essential for the generation of HCMV-permissive Allo-MDM identified gamma interferon (IFN-gamma) but not interleukin-1 or -2, tumor necrosis factor alpha, or granulocyte-macrophage colony-stimulating factor as critical components in the generation of these macrophages. In addition, although IFN-gamma was crucial for reactivation of latent HCMV, addition of IFN-gamma to unstimulated macrophage cultures was insufficient to reactivate virus. Thus, this study characterizes two distinct monocyte-derived cell types which can be distinguished by their ability to reactivate and support HCMV replication and identifies the critical importance of IFN-gamma in the reactivation of HCMV.

Cell Differentiation↗
PubMed2001

The use of sacred story in a psychiatry spirituality group.

Describes the philosophy and methodology for using sacred stories from several religious traditions with psychiatric patients. Notes how chaplains are integrated into a psychiatric unit's interdisciplinary team. Details how sacred stories are selected for use in a spirituality group, how patients are screened for participation, and how the group is facilitated. Demonstrates the benefit and value patients and staff derive from a spirituality group on a psychiatric unit.

Anecdotes as Topic↗
PubMed1999

Human cytomegalovirus infection of caco-2 cells occurs at the basolateral membrane and is differentiation state dependent.

Epithelial cells are known to be a major target for human cytomegalovirus (HCMV) infection; however, the analysis of virus-cell interactions has been difficult to approach due to the lack of in vitro models. In this study, we established a polarized epithelial cell model using a colon epithelial cell-derived cell line (Caco-2) that is susceptible to HCMV infection at early stages of cellular differentiation. Infection of polarized cells was restricted to the basolateral surface whereas virus was released apically, which was consistent with the apical and not basolateral surface localization of two essential viral glycoproteins, gB and gH. HCMV infection resulted in the development of a cytopathology characteristic of HCMV infection of colon epithelium in vivo, and infection did not spread from cell to cell. The inability of HCMV to infect Caco-2 cells at late stages of differentiation was due to a restriction at the level of viral entry and was consistent with the sequestration of a cellular receptor for HCMV. These observations provide the first evidence that restriction of HCMV replication in epithelial cells is due to a receptor-mediated phenomenon.

Caco-2 Cells↗
PubMed1999

Long-term infection and transformation of dermal microvascular endothelial cells by human herpesvirus 8.

Human herpesvirus 8 (HHV8) infects Kaposi's sarcoma (KS) spindle cells in situ, as well as the lesional endothelial cells considered to be spindle cell precursors. The HHV8 genome contains several oncogenes, suggesting that infection of endothelial and spindle cells could induce cellular transformation and tumorigenesis and promote the formation of KS lesions. To investigate the potential of HHV8 infection of endothelial cells to contribute to the development of KS, we have developed an in vitro model utilizing dermal microvascular endothelial cells that support significant HHV8 infection. In contrast to existing in vitro systems used to study HHV8 pathogenesis, the majority of dermal endothelial cells are infected with HHV8 and the viral genome is maintained indefinitely. Infection is predominantly latent, with a small percentage of cells supporting lytic replication, and latency is responsive to lytic induction stimuli. Infected endothelial cells develop a spindle shape resembling that of KS lesional cells and show characteristics of a transformed phenotype, including loss of contact inhibition and acquisition of anchorage-independent growth. These results describe a relevant model system in which to study virus-host interactions in vitro and demonstrate the ability of HHV8 to induce phenotypic changes in infected endothelial cells that resemble characteristics of KS spindle cells in vivo. Thus, our results are consistent with a direct role for HHV8 in the pathogenesis of KS.

Agar↗
PubMed1997

A guide to finding and evaluating best practices health care information on the Internet: the truth is out there?

BACKGROUND: The advent of virtually free Internet access has opened large vistas of health care information to those willing to invest a small amount of time and energy learning how to perform searches using browser software. Health care providers, organizations, and professional associations, among many others, publish "best practices" information for both administrative and clinical audiences, making these recommendations among the fastest-growing types of health care information appearing on the World Wide Web. The problem is how to find best practices among the wealth of resources on the Internet and then how to separate the proverbial wheat from the chaff. WHO IS SEEKING BEST PRACTICES ON THE INTERNET? Best practice describes a process or technique whose employment results in improved patient and/or organizational outcomes. Health care providers, managed care organizations, administrators, payers, and policy analysts are all interested in improving the quality of health care and are likely to be customers of best practices informational resources. HOW TO EVALUATE THE QUALITY OF BEST PRACTICES INFORMATION? Once the information is available on the Internet, the problem for the searcher shifts from one of quantity to quality. The best practices information seeker should stop and ask a number of questions about the quality of information, its sources, and the methods used to obtain it. CONCLUSION: The "truth" may be out there some-where in cyberspace, but locating best practices information and evaluating its quality require new skills and patience and time to practice and develop them to the point of efficiency.

Authorship↗
PubMed1996

The enhancer domain of the human cytomegalovirus major immediate-early promoter determines cell type-specific expression in transgenic mice.

The human cytomegalovirus (HCMV) major immediate-early promoter (MIEP) is one of the first promoters to activate upon infection. To examine HCMV MIEP tissue-specific expression, transgenic mice were established containing the lacZ gene regulated by the MIEP (nucleotides -670 to +54). In the transgenic mice, lacZ expression was demonstrated in 19 of 29 tissues tested by histochemical and immunochemical analyses. These tissues included brain, eye, spinal cord, esophagus, stomach, pancreas, kidney, bladder, testis, ovary, spleen, salivary gland, thymus, bone marrow, skin, cartilage, and cardiac, striated and smooth muscles. Although expression was observed in multiple organs, promoter activity was restricted to specific cell types. The cell types which demonstrated HCMV MIEP expression included retinal cells of the eye, ductile cells of the salivary gland, exocrine cells of the pancreas, mucosal cells of the stomach and intestine, neuronal cells of the brain, muscle fibers, thecal cells of the corpus luteum, and Leydig and sperm cells of the testis. These observations indicate that the HCMV MIEP is not a pan-specific promoter and that the majority of expressing tissues correlate with tissues naturally infected by the virus in the human host.

Animals↗
PubMed1996

Developmental analysis of the cytomegalovirus enhancer in transgenic animals.

The major immediate-early promoter (MIEP) of human, cytomegalovirus (HCMV) constitutes a primary genetic switch for viral activation. In this study, regulation of the enhancer-containing segment (nucleotides -670 to +54) of the HCMV MIEP attached to the 1acZ reporter gene was examined in the developing embryos of transgenic mice to identify temporal and tissue-specific expression. We find that the transgene reporter is first detected as a dorsal stripe of expression in the neural folds of embryos at day 8.5 postcoitum (p.c.). A broad expression pattern is exhibited in embryos at day 9.5 p.c. This pattern becomes more restricted by day 10.5 p.c. as organogenesis progresses. By day 14.5 p.c., prominent expression is observed in a subpopulation of central nervous system cells and spinal ganglia, endothelial cells, muscle, skin, thyroid, parathyroid, kidney, lung, liver, and gut cells, and the pancreas and submandibular and pituitary glands. This distribution pattern is discussed in relation to human congenital HCMV infection. These results suggest that the transcriptional activity of the HCMV MIEP may determine in part, the ability of the virus to specifically target developing fetal tissues in utero.

Animals↗
PubMed1994

Isolation and characterization of adenovirus-associated VA RNAs of human adenovirus type 7.

Two genes for virus-associated VA RNAs in the human adenovirus type 7 (Ad7) genome are compared to Ad2, Ad4, Ad5, simian VA RNAs, and pol III transcripts of Epstein-Barr virus. The newly identified VA RNA-encoding genes of Ad7 contain two relatively conserved intragenic promoter (elements A and B), the double-stranded RNA-dependent protein kinase (PK) active site-binding domain and transcriptional terminators. The conserved features extend to the VA RNA genes of simian and avian origin.

Adenoviruses, Human↗
PubMed1994

Restoration of telomeres in human papillomavirus-immortalized human anogenital epithelial cells.

Loss of telomeres has been hypothesized to be important in cellular senescence and may play a role in carcinogenesis. In this study, we have measured telomere length in association with the immortalization and transformation of human cervical and foreskin epithelial cells by the human papillomavirus type 16 or 18 E6 and E7 open reading frames. By using a telomeric TTAGGG repeat probe, it was shown that the telomeres of precrisis normal and E6-, E7-, and E6/E7-expressing cells gradually shortened with passaging (30 to 100 bp per population doubling). Cells that expressed both E6 and E7 went through a crisis period and gave rise to immortalized lines. In contrast to precrisis cells, E6/E7-immortalized cells generally showed an increase in telomere length as they were passaged in culture, with some later passage lines having telomeres that were similar to or longer than the earliest-passage precrisis cells examined. No consistent association could be made between telomere length and tumorigenicity of cells in nude mice. However, of the three cell lines that grew in vivo, two had long telomeres, thus arguing against the hypothesis that cancer cells favor shortened telomeres. Our results indicate that arrest of telomere shortening may be important in human papillomavirus-associated immortalization and that restoration of telomere length may be advantageous to cells with regard to their ability to proliferate.

Animals↗
PubMed1993

Interaction of HPV-18 and nitrosomethylurea in the induction of squamous cell carcinoma.

A strong correlation exists between the presence of specific types of human papillomavirus (HPV) and the development of anogenital cancer, as well as significant epidemiologic evidence suggesting smokers are at increased risk of developing cervical, vulvar and/or anal carcinomas. Primary and human papillomavirus type 18 (HPV-8)-immortalized human keratinocytes were used to address the co-carcinogenic potential of HPV and nitrosomethylurea (NMU) in tumorigenesis. Only cells containing HPV-18 and treated with NMU and the tumor-promoting phorbol ester, TPA, were transformed to a malignant phenotype. An in vitro system is described which initiates studies involving the mechanisms of HPV and chemical carcinogen co-operation in the etiology of squamous cell carcinomas.

Animals↗
PubMed1993

Alteration of the DCC tumor-suppressor gene in tumorigenic HPV-18 immortalized human keratinocytes transformed by nitrosomethylurea.

A human papillomavirus type 18 (HPV-18)-immortalized human keratinocyte cell line (1811) has been transformed to tumorigenicity in nude mice by treatment with the carcinogen nitrosomethylurea (NMU). The NMU transformants (1811-NMU-T) showed additional chromosome alterations as compared with parental 1811 cells, including 18q deletion in two of two 1811-NMU-T lines analysed. Restriction fragment length polymorphism (RFLP) analysis indicated that both 1811-NMU-T lines had lost one allele of the 18q deleted in colon cancer (DCC) tumor-suppressor gene. Reverse transcriptase polymerase chain reaction (RT-PCR) showed that DCC expression was absent or barely detectable in the 1811-NMU-T cells as compared with 1811 or normal keratinocytes, suggesting that the remaining DCC allele in the 1811-NMU-T cells was also altered. These studies indicate that reduction or loss of DCC expression may be an important step in NMU transformation of HPV-immortalized cells to tumorigenicity.

Adenomatous Polyposis Coli↗
PubMed1992

Degradation of aliphatic halogen-substituted pesticides by dehalogenase isolated from Pseudomonas alcaligenes. Identification and properties of the enzyme.

Some characteristics of a 2,2-dichloropropionate dehalogenase induced in a bacterial strain capable of degrading high concentrations of the herbicide dalapon were studied. Polyacrilamide gel electrophoresis of the crude cell free extracts identified only one type of dehalogenase. The single enzymatic protein showed activity against a variety of chlorinated aliphatic acids but differed in their activity levels. Thus activity in mumol substrate converted (mg protein)-1 min-1 was 2-monochloropropionate 0.65, 2,2-dichloropropionate 0.56, 2-monochloroacetate 1.70 and 2,2-dichloroacetate 1.00. In the crude extracts, the enzyme activity against 2,2-dichloropropionate was optimal at a broad pH range with a mid-point at pH 9.5 and apparent Km values were within the range 0.23-0.73 mM.

Biodegradation, Environmental↗
PubMed1992

Immortalization of primary human smooth muscle cells.

Primary human aortic and myometrial smooth muscle cells (SMCs) were immortalized using an amphotropic recombinant retroviral construct containing the E6 and E7 open reading frames (ORFs) of human papillomavirus type 16. The SMCs expressing the E6/E7 ORFs have considerably elevated growth rates when compared with nonimmortalized control cells and show no signs of senescence with long-term passage. The first SMC line derived in this study has been maintained in continuous tissue culture for greater than 1 year (greater than 180 population doublings). The immortalized SMCs have decreased cell size and decreased content of muscle-specific alpha-actin filaments as determined by indirect immunofluorescence. Southern blot analysis has demonstrated the stable integration of the E6/E7 ORFs in the retrovirally infected cells, and radioimmunoprecipitation has confirmed the continued expression of the E6 and E7 genes. Cytogenetic studies of the SMC lines have revealed essentially diploid populations except for the myometrial clonal line, which became aneuploid at late passage (greater than 125 doublings). These cell lines were not tumorigenic in nude mice.

Animals↗
PubMed1992

Viral integration and fragile sites in human papillomavirus-immortalized human keratinocyte cell lines.

Human papillomavirus (HPV) types 16 and 18 integration sites were mapped in six HPV-immortalized human keratinocyte cell lines by fluorescence in situ hybridization (FISH). Mapping of HPV sequences in these cell lines revealed that HPV integration varied in copy number and location but that integration sites were stable over extended passages in culture. Integration occurred at different sites throughout the genome and did not correspond to the location of specific cellular genes. However, integration sites were consistent with integration near or within known fragile sites in five of the six cell lines. Induction of aphidicolin-sensitive fragile sites in one cell line prior to in situ hybridization revealed that integrated HPV DNA was disrupted by fragile-site expression, suggesting that integration occurred within a fragile site.

Cell Line, Transformed↗
PubMed1991

Progression of human papillomavirus type 18-immortalized human keratinocytes to a malignant phenotype.

We have developed a model system for progression of human epithelial cells to malignancy, using a human papillomavirus type 18 (HPV-18)-immortalized human keratinocyte cell line. Cells of cell line FEP-1811 were nontumorigenic in athymic mice through at least 12 passages in culture, but after 32 passages were weakly tumorigenic, producing tumors that regressed. After 62 passages they produced invasive squamous cell carcinomas that grew progressively. The progression to malignancy was associated with an increase in the efficiency of forming colonies in soft agar and with altered differentiation properties. In an organotypic culture system, FEP-1811 cells at passages 12 and 32 exhibited features typical of premalignant intraepithelial neoplasia in vivo, and cells at passage 68 exhibited features consistent with squamous cell carcinomas. No change in copy number of the transfected HPV-18 genome or in the level of expression of the viral transforming gene products E6 and E7 was detected between tumorigenic and nontumorigenic cells. Cytogenetic analysis of cells at early, middle, and late passage levels and cells cultured from tumors revealed that several chromosomal abnormalities segregated with the tumorigenic cell populations.

Animals↗