http://biochem.dental.upenn.edu/ (World Wide Web Directory, 06/1995)
The Center for Oral Health Research (COHR) was constituted in 1970 as the basic sciences research core of the Univrsity of Pennsylvania School of Dental Medicine. The University, which is a privately endowed nonsectarian institution founded by Benjamin Franklin in 1740, has played a significant role in the development of contemporary ideas in both the humanities and sciences. Currently, there are 12,000 undergraduate and 9,000 graduate and professional students.
The major mission of COHR is to cultivate biomedical and oral health relaed research activities. The Center recives considerable NIH support and is a major national site for oral and dental research. There is extensive collaboration with scientists in other schools of the University as well as with those of other institutions. COHR is located adjacnt to the School of Dental Medicine in the Leon of Veterinary Medicine, three blocks from the Medical School and within easy walking distance of the Hospital of the University of Pennsylvania and the Childrens Hospital of Philadelphia. For twenty years, COHR has trained undergraduate and graduate students as well as postdoctoral fellows and visitng scientists.
This brochure describes the ongoing scientific studies of infection and the host response, bone, tooth and extracellular matrix, invasive pathogens and viral infection. Core support facilities include resources for protein analysis, fluorescence- activated cell sorting and protein/peptide structure modeling as well as a Clinical Research Center with its Oral Fluids Laboratory.
Scientists in COHR welcome inquiries from students and graduates in the life sciences concerning opportunities to participate in ongoing research.
Investigators are exploring the inflammatory response of neutrophils to bacteria. These studies have identified a number of host and bacterial derived factors which influence the ability of bacteria to cause disease. Oral bacteria can produce a potent toxin which specifically kills neutrophils and increases the infectivity of the microorganism. The gene for this leukotoxin has been identified, cloned and expressed in E. coli. Investigators are using a molecular biological approach to further dissect the molecule and understand its mechanism of action.
Another molecule isolated from oral flora is involved in immunoregulation. It suppresses the immune response by activating T-suppressor cells. Potentially, this factor could be used therapeutically as an immunosuppressive agent. The host response to spirochetes is also a topic of interest, and investigators are looking at the mechanism whereby these organisms evade the host defense system.
Human saliva modulates the colonization of bacteria, and also interferes with HIV infectivity. Investigators at the COHR are currently studying the interaction between salivary proteins and a variety of bacteria and viurses. Particular attention is focused on the ability of the host to neutralize virulence factors such as adhesins, antibodies, and agglutinins. Studies are also being conducted into the effect of heavy metals on blood cells with the objective of determining whether elements in dental amalgam may be injurious to leukocyte function.
Cartilage and Bone Development: The skeletal system of vertebrates, consisting of cartilage and bone, undergoes a complex developmental pathway which is characterized by profound changes both in the composition of the extracellular matrix and in energy metabolism. During this developmental process, mesenchymal cells differentiate into chondrocytes, the cells that produce cartilage. In endochondral growth cartilage, a structure that mediates bone growth, chondrocytes mature and produce a mineralized matrix. Ultimately this matrix is replaced by mineralized bone. The cellular and molecular mechanisms mediating this developmental process are the subject of an extensive COHR research program.
COHR scientists are probing the molecular mechanisms regulating changes in collagen types during maturation of cells which form bone and cartilage, as well as those controlling the induction of a unique protein, type X collagen, during cartilage maturation.
Experiments are being performed to characterize the structure- function relationships of alkaline phosphatase, an enzyme which is expressed at high levels in all mineralizing tissues, yet whose role in the calcification process is enigmatic. Related studies are aimed at elucidating the metabolic status of the cells of the growth plate and relating changes in gene expression to oxidative metabolism, cell maturation and calcification.
Additional investigations are directed at exploring the mechanism of matrix removal by osteoclasts. Model systems have been developed for studying the mechanism of cleft palate formation and evaluating the interaction between bone cells and implanted ceramics and metals. Futher, extracellular structural proteins involved in mineral deposition are being investigated to by two- dimensional NMR spectroscopy. Other protein substrate interactions are being analyzed by 13C and 15N nuclear magnetic resonance (NMR) spectroscopy.
Enamel: Tooth enamel is the hardest tissue in the human body. Its unique class of matrix proteins, amelogenins, participate in mineral formation. Center investigators are exploring the structure and function of these proteins by cloning the DNA coding sequences into plasmids for expression in E. coli. Transgenic mice are being used to study the effects of gene alteration in vivo and to create an animal model for the human enamel defect hereditary amelogenesis imperfecta.
Elastin: Tissues which require flxibility for function produce elastin as an extracellular matrix protein. Molecular mechanisms for expression of the elastin gene are currently being identified. Research workers in the Levy Center are dissecting the mechanism by which the tropoelastin precursor interacts with itself and accessory proteins to from mature elastin fibers. In addition, clinical testss are being devised to monitor patients with emphysema, in which lung elasticity is compromised during disease grogression and treatment.
Ongoing research efforts in COHR are focused on the application of genetic techniques to detect and characterize periodontal pathogens. DNA probes are used for diagnosis of oral pathogens and to differentiate strains of periodontal bacteria obtained from epidemiological studies. Other studies are aimed at determining the molecular basis of bacterial adherence. Emphasis is placed on the isolation and characterization of adhesins from Streptococcus sanguis, pili from Actinobacillus actinomycetemcomitans, and intergeneric coaggregation receptors from Fusobacterium nucleatum. Work is also progressing towards the elucidation of the molecular mechanisms of pathogenicity of periodontopathic bacteria. Cytotoxic macromolecules as well as extrachromosomal genetic elements, such as plasmids and bacteriophages, produced by these bacteria are being isolated and characterized.
The research effort is directed towards understanding the etiology and diagnosis of periodontal diseases. Can bacterial components and products be used to construct diagnostic, genetic or immunologic probes, which can detect the earliest presence of specific pathogenic strains? Can the use of these probes provide new information concerning the epidemiology of oral diseases? What are the mechanisms that allow diverse bacterial species to become established in the oral cavity? Understanding the molecular basis of bacterial adherence could lead to the development of agents that inhibit colonization. COHR scientists are determining which specific bacterial components or products have the potential to contribute to the development of oral infectious diseases. Ultimately, it may be possible to selectively eliminate, at or after the onset of infection, those species of oral bacteria that promote the disease state.
Four glycoproteins, gC, gE, gG and gI, may play a significant role in pathogenesis. The glycoproteins gE and gI function as Fc receptors for human IgG; gC is a receptor for complement components C3b and iC3b and may modulate complement mediated host responses. At present, ongoing studies concentrate on gC and gD.
Recent studies have shown that gD interacts with a specific cell receptor; current and future experiments are being directed at this receptor interaction and its importance in the entry process. In addition, gD is being tested as a potential human subunite vaccine. To map the portion of gD which is critical for its function, researchers COHR have used site-directed mutagenesis techniques to create mutations in the gD gene. Other studies are focused on the effects of those mutations, using immunological, as well as functional assays. These studies may also provide information about the folding of gD into its native structure.
Experiments are in progress to identify the interaction of gC with the complement of component C3b. Thus far, several regions of gC-1 and gC-2 have been implicated in C3b receptor activity and one of these regions shows homology with a portion of the human complement receptor CRI. The goal is to evaluate the role of the glycoproteins in HSV infection adn pathogenesis in order to design a paradigm for intervention.
Ultrastructural and Imaging Analysis Core Facility: The facility prepares bological tissues for most types of ultrastructural studies. The facility contains a JEOL 100 CXII transmission electron microscope and a JEOL 330 A scanning electron microscope equipped with a KEVEX energy dispersive X-ray microanalysis unit. Routine analysis is provided as a service while more extensive work usually involves collaborative arrangements with the Director, Dr. Peter Berthold.
Immunology Core Facility: Under the supervision of Dr. Edward Lally, polyclonal and monoclonal antibodies have been developed against a wide variety of antigens including leukotoxin, human collagen types III and IV, DNP Herpes simplex glycoprotein D, salivary bacterial agglutinin, enamel proteins, and elastin.
Protein Analytical Laboratory: Under the direction of Dr. Bill Abrams, this facility provides structural information related to proteins or peptides of interest. Proteins are routinely characterized by amino acid composition determination as well as by automated protein/peptide N-terminal microsequencing and manual C-terminal microsequencing. The level of material required for these analyses is in the low pmole range. The facility utilizes a Miligen/Biosearch 6600 ProSequencer with on- line PTH amino acd chromatographic separation and automated identification and quantification of residues. Amino acid composition is performed by precolumn derivitization of protein hydrolysates using reverse phase separation of PTC-amino acids. A Water 625LC is available for peptide mapping and isolation of 100 pmole levels of fragments for sequence analysis.
Molecular Biology Core Facility: Under the supervision of Dr. Carolyn Gibson, oligonucleotides for sequencing primers and probes for Souther blots and PCR analysis are produced by a Miligen/Biosearch DNA synthezider. Investigators are provided with experitise and support for sequencing of DNA. In addition, prokaryote expression vectors are available for protein or peptide over-expression in E. coli.
Imaging and Computer Core Facility: This facility is under the directio nof Drs. Ellis Golub, Eileen Jaffe and John Haselgrove. It provides computer based DNA and protein comparative sequence analysis, modeling of protein secondary structure based upon deduced or determined primary amino acid structure; imae analysis; NMR analysis of protein structure; programming and database analysis; and computer network expertise. Data reduction and analysis is facilitated by linkage to information data bases and a Silicon Graphics personal Iris work station. Flow Cytometry and Cell Sorter Laboratory: This facility, directed by Dr. Bruce Shenker, has been developed for the purpose of analyzing and sorting cells and chromosomes. It is equipped with a Becton Dickinson FACStar Plus flow cytometer and dual lasers. This technology includes six parameter analyses (four color fluorescence), light scattering, calcium imaging and cell cycling. The facility utilizes state-of-the-art computer analysis and presentation graphics.
FOR MORE INFORMATION ABOUT COHR COMMUNICATE WITH:
Dr. Joel Rosenbloom, Associate Dean for Research, School of Dental Medicine, Leon Levy Center for Oral Health Research, University of Pennsylvania, 4001 Spruce Street, Philadelphia, PA 19104-6003.
Telephone: 215-898-8994 FAX: 215-898-5243
Robert Hooper
robert@biochem.dental.upenn.edu