Molecular Immunology Group
Martin J. Hessner, PhD
The development of complex diseases involves the interplay of multiple genes, cell types and environmental factors. An integrated perspective of how these factors interact is vital towards the dissection and understanding of complex disease as well as a prerequisite for the development of successful treatment and prevention strategies. We have recognized the potential of gene expression profiling in systems biology as well as its technical and analytical complexities. Therefore, our laboratory has focused on the development of a cost-effective, highly-controlled three-color array platform focused on the acquisition of quality data [1-9] (Figure 1).
We have applied functional genomics to the events leading to type 1 diabetes mellitus (T1DM), a complex disease that targets the insulin-producing pancreatic ß cells. We use the nondiabetic Wistar-Furth (WF) rat and congenic BioBreeding (BB) rat derivatives. BB DRlyp/lyp rats are spontaneously diabetic due to a mutation in the Gimap5 gene [10], rendering them deficient in regulatory T (TREG) cells [11-13]. In contrast, DR+/+ rats possess an intact Gimap5 gene and do not develop spontaneous T1DM, but become diabetic upon depletion of TREG cells. T1DM in BB rats also involves cytotoxic T cells, since their depletion abrogates disease [14]. Through expression profiling of pancreatic lymph nodes (PLN) and isolated PLN subpopulations [15, 16], we identified a key role for mast cells in BB rat diabetogenesis. We have confirmed their importance by delaying/preventing T1DM in BB rats with two different mast cell inhibitors [16]. Furthermore, unlike the WF, we find the congenic BB strains predisposed to T1DM in that their pancreatic ß cells begin expressing mast cell recruiting cytokines early in life (Figure 2). These findings suggest that mast cells may not only precipitate islet destruction, but may serve as potential initiators of the adaptive T cell mediated response. Our results are consistent with human and rodent studies implicating mast cells in the initiation and progression of autoimmunity [17]. Our efforts continue to better understand the events leading to T1DM by employing genomics, histological, immunological, and bioinformatics strategies to the analysis of the BB rat.
Understanding the active proinflammatory mechanisms at and prior to type 1 diabetes mellitus onset is hindered in humans, given that relevant tissues are inaccessible and pancreatic immune responses are dilute and difficult to measure in the periphery by traditional approaches. Thus, our human studies have used a sensitive array based genomics bioassay where the serum of recent onset T1DM subjects is used to induce gene expression in healthy peripheral blood mononuclear cells (PBMCs). We find that serum of recent-onset diabetics induce a unique pro-inflammatory gene expression signature that includes numerous innate immunity genes and genes regulated by IL-1ß. The signature is completely distinct from that induced by normal control sera. Remarkably, this molecular signature is evident >5 years prior to T1DM onset and holds value in predicting onset in "at-risk" subjects and as a potential measure in primary and secondary prevention trials.
 |
| Figure 1. Hybridization of d40 and d65 DRlyp/lyp directly labeled total thymus RNA targets to 36k cDNA arrays. A. Prehybridization fluorescein image of "A" array (1st 18,000 probes). Note heterogeneity in spot intensity due to differing PCR efficiencies, and spotting buffer negative controls (lower right corner of inset). B. Post hybridization cyanine images of reverse-labeled replicates (slides 16 and 22: d40 Cy5, d65 Cy3; slides 12 and 14 d40 Cy3, d65 Cy5). Circled on each image is the probe specific for sterol coenzyme A destaturase; 6-fold over-expressed at d40 relative to d65. Dilution series for Arabidopsis rubisco activase (left) and Nac1 (right) spiked into the Cy5 and Cy3 labeling reactions at 1:30 and 30:1, respectively, are shown. |
|
 |
|
Figure 2. Immunofluorescent staining of pancreata. Columns A and B, staining of WF and DR+/+ frozen sections with anti-glucagon (FITC, green) and anti-eotxin (texas red), respectively. Columns C and D, staining of WF and DR+/+ with anti-insulin (FITC, green) and anti-eotxin (texas red), respectively. Time points d20 through d60 are indicated by row, bottom row lacks primary anti-eotaxin antibody as a negative control. All imaging used 40X dry objective lens.
|
|
|
|
| Figure 3. Profiles of 192 unique UniGenes significantly regulated (|fold change|>1.5; Student's t-test p<0.01) after healthy PBMCs are cultured with recent-onset (RO, n=12) versus healthy control (HC, n=12) sera. All data was normalized with that of the autologous induction to account for gene expression induced by placing PBMCs into culture. Thus data is represented as a ration of HC:auto or RO:auto. |
|
Journal Publications/Original Papers:
Geoffrey, R., S. Jia, A.E. Kwitek, J., S. Ghosh, Å. Lernmark, X. Wang, M.J. Hessner. 2006. Evidence of a functional role for mast cells in the development of type 1 diabetes mellitus in the BioBreeding rat. The Journal of Immunology 177(10):7275-86.
Wang X., S. Jia, L. Meyer, B. Xiang, L.Y. Chen LY, N. Jiang, C. Moreno, H.J. Jacob, S. Ghosh, M.J. Hessner. 2006. Comprehensive quality control utilizing the prehybridization third-dye image leads to accurate gene expression measurements by cDNA microarrays. BMC Bioinformatics 7:378.
Hessner, M.J., M. Liang, A.E. Kwitek. 2006. The Application of Microarray Analysis to Pediatric Diseases. Pediatric Clinics of North America. 53:579–590.
Hessner, M.J., B. Xiang, S. Jia, R. Geoffrey, S. Holmes, L. Meyer, S. Muheisen, X. Wang. 2006. Three-color cDNA microarrays with prehybridization quality control yield gene expression data comparable to that of commercial platforms. Physiological Genomics 25:166-78
Liang, M., A.W. Cowley Jr., M.J. Hessner, J. Lazar, D.P. Basile, J.L. Pietrusz. 2005. Transcriptome analysis and kidney research: Toward systems biology. Kidney International 67:2114-22.
Yang, J., X. Wang, J. Moibi, M.J. Hessner, S. Greene, J. Wu, R.K. Wong, S. Sukumvanich, B.A. Wolf, Z.Gao 2004. Leucine culture reveals that ATP synthase functions as a fuel sensor in pancreatic β cells. Journal of Biological Chemistry. 279(52):53915-23.
Hessner, M.J., X. Wang, L. Meyer, R. Geoffrey, S. Jia, J. Fuller, A. Lernmark, S. Ghosh. 2004. Involvement of eotaxin, eosinophils, and pancreatic predisposition in development of type 1 diabetes mellitus in the BioBreeding rat. The Journal of Immunology, 173(11):6993-7002.
Schlict, M., B. Matysiak, T. Brodzeller, X. Wen, H. Liu, G. Zhou, R. Dhir, M.J. Hessner, P. Tonellato, M. Suckow, M. Pollard, M.W. Datta. 2004. Cross-species global and subset gene expression profiling identifies genes involved in prostate cancer response to selenium. BMC Genomics. 5(1)58.
Hessner, M.J., L. Meyer, J. Tackes, S. Muheisen, X. Wang. 2004. Immobilized probe and glass surface chemistry as variables in microarray fabrication. BMC Genomics, 5(1):53.
Morrison, J., K. Knoll, M.J. Hessner, M. Liang. 2004. Effect of high glucose on gene expression in mesangial cells: Up-regulation of the thiol pathway is an adaptational response. Physiological Genomics, 17(3):271-82.
Zhou, G., X. We, H. Liu, M. Schlicht, M.J. Hessner, P.J. Tonellato, M.W. Datta. 2004. BEAR GeneInfo: A tool for identifying gene-related biomedical publications through user modifiable queries. BMC Bioinformatics, 5:46.
Patnaik, M., J. Dlott, R. Fontaine, M.J. Hessner, K. Joyner, M.R. Ledford, E. Lau, C. Moehlenkamp, J. Amos-Wilson, B. Zhang, T. Williams. 2004. Detection of genomic polymorphisms associated with venous thrombosis using the InvaderÒ biplex assay. Journal of Molecular Diagnostics, 2:137-144.
Hessner, M.J., V.K. Singh, X. Wang, S. Khan, M.R. Tschannen, T.C. Zahrt. 2004. Utilization of a labeled tracking oligonucleotide for visualization and quality control of spotted 70-mer arrays. Genomics, 5:12.
Wang, X., N. Jiang, X. Feng, Y. Xie, P. Tonellato, S. Ghosh, M.J. Hessner. 2003. A novel approach for microarray image processing using third-dye array visualization technology. IEEE Transactions on Nanobioscience, 2:103-201.
Waukau, J., S. Jailwala, Y. Wang, H-J. Khoo, S. Ghosh, X. Wang, M.J. Hessner. 2003. The design of a gene chip for functional immunological studies on a high-quality control platform. Annals of the New York Academy of Sciences, 1005: 284-287.
Pati, N., S. Ghosh, M.J. Hessner, H-J Khoo, X. Wang. 2003. Difference in gene expression profiles between human CD4+CD25+ and CD4+CD25- T cells. Annals of the New York Academy of Sciences, 1005: 279-283.
Wang, X., M.J. Hessner, Y. Wu, N. Pati, S. Ghosh. 2003. Quantitative quality control in microarray experiments and the application in data filtering, normalization, and false-positive rate prediction. Bioinformatics, 19:1341-1347.
Hessner, M.J., X. Wang, S. Khan, L. Meyer, M. Schlicht, J. Tackes, M.W. Datta, H. J. Jacob, S. Ghosh. 2003. Use of a three-color cDNA microarray platform to measure and control support-bound probe for improved data quality and reproducibility. Nucleic Acids Research, 31:e60.
Hessner M.J., X. Wang, K. Hulse, L. Meyer, Y. Wu, S. Nye, S.W. Guo, S Ghosh. 2003. Three color cDNA microarrays: Quantitative assessment through the use of fluorescein-labeled probes. Nucleic Acids Research, 31:e14.
Ekman, G.C., R. Billingsly, M.J. Hessner. 2002. Rh genotyping: Avoiding false-negative and false-positive results among individuals of African ancestry. American Journal of Hematology 69:34-40.
Raife, T.J., S.R. Lentz, B. Atkinson, S.K. Vesely, M.J. Hessner. 2002. Factor V Leiden: a genetic risk factor for thrombotic microangiopathy in patients with normal von Willebrand factor-cleaving protease activity. Blood 99(2):437-42.
Hessner, M.J., K.D. Friedman, K. Voelkerding, S. Huber, D. Ryan, B. Nucci, M. Ledford. 2001. A multisite study for genotyping of the factor II (Prothrombin) G20210A mutation by the Invader assay. Clinical Chemistry 47:2048-2050.
Hessner, M.J., D.M. Dinauer, R. Kwiatkowski, B. Neri, T.J. Raife. 2001. Measurement of vascular disease-associated polymorphisms among age-stratified volunteer blood donors. Clinical Chemistry 47:1879-1884.
Lazzaro B., A. Anderson, A. Kadacszy-Balla, M.J. Hessner. 2001. Antigenic characterization of medullary carcinoma of the breast:HLA-DR expression in lymph node positive cases. Applied Immunohistochemistry and Molecular Morphology 9:234-241.
Hessner, M.J., Luhm, R.A. 2000. The C536T transition in the tissue factor pathway inhibitor (TFPI) gene does not contribute to risk of venous thrombosis among carriers of factor V Leiden. Thrombosis and Haemostasis 84:424-425.
Curtis, B.R., J.T. Edwards, M.J. Hessner, J.P. Klein, R.H. Aster. 2000. Blood group A and B antigens are strongly expressed on platelets of some individuals. Blood 96:1574-1581.
Hessner, M.J., M.A. Budish, K.D. Friedman. 2000. Genotyping of factor V G1691A (Leiden) without the use of PCR by invasive cleavage of oligonucleotide probes. Clinical Chemistry 46:1051-1056.
Ekman, G.C., M.J. Hessner. 2000. Screening of six racial groups for the intron 5 G→A 3' splice acceptor mutation responsible for the Polynesian Kidd (a-b-) phenotype: the null mutation is not always associated with the JKB allele. Transfusion 40:888-889.
Luhm, R.A., D.B. Bellissimo, A.J. Uzgiris, W.R. Drobyski, M.J. Hessner. 2000. Quantitative evaluation of post- bone marrow transplant engraftment status using fluorescently-labeled variable number of tandem repeats (VNTRs). Molecular Diagnosis 5:129-138.
Ledford, M., K.D. Friedman, M.J. Hessner, C. Moehlenkamp, T.M. Williams, R. Larson. 2000. A multi-site study for detection of the factor V (Leidenmutation from genomic DNA using a homogeneous InvaderTM microtiter plate fluorescence resonance energy transfer (FRET) assay. Journal of Molecular Diagnostics 2:97-104.
Dinauer, D.M., R.A. Luhm, A.J. Uzgiris, D.D. Eckels, M.J. Hessner. 2000. Sequence-based typing of HLA class II DQB1. Tissue Antigens. 55:364-368.
Dinauer, D.M., K.D. Friedman, M.J. Hessner. 1999. Allelic distribution of the glycoprotein Ia (2-Integrin) C807T/G873A dimorphisms among Caucasian venous thrombosis patients and six racial groups. The British Journal of Haematology. 107:563-565.
Hessner, M.J., D.M. Dinauer, R.A. Luhm, J.L. Endres, R.R. Montgomery, K.D. Friedman. 1999. The prothrombin 20210GA genotypes acts as a cooperative risk factor for venous thrombosis among factor V 1691GA (Leiden) carriers. The British Journal of Haematology. 106:237-239.
Drobyski, W.R., M.J. Hessner, J. Klein, C. Kabler-Babbit, D.H. Vesole, C.A. Keever-Taylor. 1999. T-cell depletion plus salvage immunotherapy with donor leukocyte infusions as a strategy to treat chronic phase chronic myelogenous leukemia patients undergoing HLA-identical sibling or partially matched family donor transplants. Blood. 94:434-441.
Hessner, M.J., R.A. Luhm, S.L. Pearson, D.J. Endean, K.D. Friedman, R.R. Montgomery. 1999. Prevalence of prothrombin G20210A, factor V G1691A (Leiden), and methylene tetrahydrofolate reductase C677T in seven different populations by multiplex allele-specific PCR. Thrombosis and Haemostasis. 81:733-738.
Hessner, M.J., R.A. Pircon, S.T. Johnson, R.A. Luhm. 1999. Prenatal genotyping of the Duffy blood group system by allele-specific polymerase chain reaction. Prenatal Diagnosis. 19:41-45.
Hessner, M.J., S. Shivaram, D.M. Dinauer, B.R. Curtis, D.J. Endean, R.H. Aster. 1999. Neutrophil antigen (Fc RIIIB) SH gene frequencies in six racial groups. Blood. 93:1115-1116.
Hessner, M.J., R.A. Pircon, S.T. Johnson, R.A. Luhm. 1998. Prenatal genotyping of Jka and Jkb of the human Kidd blood group system by allele-specific polymerase chain reaction. Prenatal Diagnosis. 12:1225-1231.
Drobyski, W.R., M.J. Hessner. 1998. The use of the polymerase chain reaction to predict for subsequent relapse in unrelated marrow transplantation for chronic myelogenous leukemia. Leukemia and Lymphoma. 31:317-323.
Juckett, M., P. Rowlings, M.J. Hessner, C.A. Taylor, W. Burns, B. Camitta, J.T. Casper, W.R. Drobyski, G. Hanson, M.M. Horowitz, C Lawton, D.A. Margolis, D. Peitryga, D. Vesole. 1998. T-cell depleted allogeneic bone marrow transplant for high risk non-Hodgkin lymphoma: clinical and molecular follow-up. Bone Marrow Transplantation. 21:893-899.
Drobyski, W.R., C. Pelz, C. Kabler-Babbitt, M.J. Hessner, L.A. Baxter-Lowe, C.A. Keever-Taylor. 1998. Successful unrelated marrow transplantation for patients over the age of 40 with chronic myelogenous leukemia. Biology of Blood and Marrow Transplantation. 4:3-12.
Pearson, S.L., M.J. Hessner. 1998. A1,2BO1,2 genotyping by multiplexed allele-specific polymerase chain reaction. The British Journal of Haematology, 100:229-234.
Hessner, M.J. 1997. Clinical application of allele-specific polymerase chain reaction genotyping for identification of fetuses at risk for immune cytopenic disorders. Biotech Lab International, 2:8-11.
Drobyski, W.R., D.J. Endean, J. Klein, M.J. Hessner. 1997. Detection of BCR/ABL transcripts using the polymerase chain reaction is highly predictive for relapse in patients transplanted with unrelated marrow grafts for chronic myelogenous leukemia. The British Journal of Haematology, 98:458-466.
Hessner, M.J., T.A. Agostini, D.B. Bellissimo, D.J. Endean, R.A. Pircon, N.E. Kirschbaum. 1997. The sensitivity of allele-specific PCR can obviate concern of maternal contamination when fetal samples are genotyped for immune cytopenic disorders. The American Journal of Obstetrics and Gynecology, 176:327-333.
Hessner, M.J., B.R. Curtis, D.J. Endean, R.H. Aster. 1996. Determination of neutrophil antigen NA gene frequencies in five different ethnic groups by the polymerase chain reaction with sequence-specific primers (PCR-SSP). Transfusion, 36:895-899.
Hessner, M.J., J.G. McFarland, D.J. Endean. 1996. Genotyping of K1 and K2 of the human Kell blood group system by the polymerase chain reaction with sequence-specific primers (PCR-SSP). Transfusion, 36:495- 499.
Hessner, M.J., D.J. Endean, J.T. Casper, M.M. Horowitz, C.A. Keever-Taylor, M.S. Roth, N. Flomenberg, W.R. Drobyski. 1995. Use of unrelated marrow grafts compensates for loss of graft-versus-leukemia reactivity after T-cell depleted allogeneic BMT for chronic myelogenous leukemia. Blood, 86:3987-3996.
Skogen, B., D.B. Bellissimo, M.J. Hessner, S. Santoso, R.H. Aster, P.J. Newman, J.G. McFarland. 1994. Rapid determination of platelet alloantigen genotypes by polymerase chain reaction using allele-specific primers. Transfusion, 34:955-960.
Hessner, M.J., M.S. Roth, W.R. Drobyski, L.A. Baxter-Lowe. 1994. Development of a sensitive, highly controlled assay for molecular detection of the Philadelphia chromosome in patients with chronic myelogenous leukemia. Genetic Analysis: Techniques and Applications, 11:90-94.
Hessner, M.J., L.A. Baxter-Lowe. 1992. Characterization of novel HLA-DPB1 alleles by oligotyping and nucleotide sequencing. Tissue Antigens, 40:261-263.
Hessner, M.J., P.J. Wejksnora, M.L.P. Collins. 1991. Construction, characterization, and complementation of Rhodospirillum rubrum puf region mutants. Journal of Bacteriology, 173:5712-5722.
BOOK CHAPTERS:
Wang, X., and M.J. Hessner. Quantitative quality control of microarray experiments: toward accurate gene expression measurements. 2006. In Cambridge University Press: Gene Expression Profiling by Microarrays - Clinical Implications. Wolf-Karsten Hofmann (editor).
Hessner, M.J. and B.R. Curtis. Prenatal genotyping for identification of fetuses at risk for immune cytopenic disorders. 2004. In: Humana Press, Molecular Diagnostics for the Clinical Laboratorian. W.B. Coleman and G.J. Tsongalis (editors).
Broeckel, U. and M.J. Hessner. 2004. Single Nucleotide Polymorphisms: Testing DNA Variation for Disease Association In: Humana Press, Molecular Diagnostics for the Clinical Laboratorian. W.B. Coleman and G.J. Tsongalis (editors).
Hessner, M.J., X. Wang, S. Ghosh. Genetics of Type 1 Diabetes. 2004 In: Lippencott, Williams and Wilkins, Diabetes Mellitus: A Fundamental and Clinical Text. D. LeRoith, S. Taylor, J. Olefsky (editors).
Hessner, M.J. and D.B. Bellissimo. Prenatal Genotyping of the RhD locus by allele-specific polymerase chain reaction for the identification of fetuses at risk for hemolytic disease of the newborn. 2001. In: Humana Press, Methods in Molecular Medicine, Molecular Pathology Protocols. A. Killeen (editor).