DIVISION OF TRANSPLANTATION PATHOLOGY

DIVISION OF TRANSPLANTATION PATHOLOGY

                                                   

Clinical Activities

Formally established in 1990, the Division of Transplant Pathology plays an integral part in the comprehensive transplant program at the University of Pittsburgh Medical Center and its satellite center in Palermo, Italy.  The mission of the Division is to provide up-to-date laboratory services emphasizing continued excellence in patient care, to provide expert consultation, to foster an academic environment devoted to education and training, and to conduct basic and clinically oriented research in transplantation pathology.  A nucleus of professionals with expertise in pathology, immunology and molecular biology is actively involved in many transplantation related research projects at the Thomas E. Starzl Transplantation Institute.

 

The Division offers a wide range of laboratory tests and tissue analyses for patients and experimental studies.  Routine histology, specialized immunocytochemistry and in situ hybridization procedures are performed on tissues from patients under consideration for transplantation, on native organ resections and on post-transplant biopsies.  These procedures assure consistency of analysis and continuity of patient care.  Post-transplant monitoring assays deal with lymphocyte reactivity, cytokine production and detection of cytomegalovirus, Epstein-Barr virus, and Hepatitis viruses, among other infectious agents.

 

The division has established and maintains an active telepathology consultation service.  We developed specific HIPAA-compliant software with Diversified Services, Inc. that is used to triage and respond to cases.  We have also recently integrated robotic microscope capabilities (Trestle Systems) into the software applications.

 

Dr. Adriana Zeevi is the Director and Dr. Alin Girnita is Assistant Director of the UPMC-Presbyterian Tissue Typing Laboratory, which offers histocompatibility-testing services to the organ transplantation program at the University of Pittsburgh Medical Center.  This laboratory performed donor workups as follows:  liver (112), living-related liver (19), heart (75), lung and heart/lung (130), kidney (112), canceled kidney donors (255) living-related kidney (78), pancreas and pancreas/kidney (26) and small bowel/multi viseral (42) for a total of 838. Two hundred fifty-five cadaveric kidney donors were worked up in the laboratory, but not used.  The laboratory also began to provide services to the VA Kidney Transplant Program, this year.  These numbers are included in the totals.

 

The Division offers comprehensive consultation in liver, kidney, and solid organ transplantation pathology to community and university-based physicians.  The scope of this service encompasses the evaluation of liver, heart, intestinal and kidney disease in prospective transplantation candidates, donor recipient histocompatibility issues, the examination of native explanted organs, and the evaluation of post-transplant biopsies or resections to evaluate the causes of allograft dysfunction such as infection, ischemia, rejection, technical complications, recurrent disease and post-transplant malignancies.  These consultation services are also available through the Internet (http://path.upmc.edu/divisions/transpath/hepa00.html).

 

For the period of July 1, 2008 through June 30, 2009, the professional workload (generated from CoPath; telepathology cases excluded) for each member was:

 

 

PHSC-TX-Red

PHSC-UP-UPMC-Yellow

PHSC-UP-Outside-

Yellow

PHSG-EM

PHSG-other

PHTX-Surg

PHTX-EM

Autopsy

Total

 

 

 

 

Demetris

37

36

493

2

9

918

2

0

1497

 

 

Nalesnik

99

13

105

4

7

957

3

0

1188

 

 

Ochoa

125

16

73

0

8

762

1

0

985

 

 

Randhawa

93

18

108

0

8

733

2

0

962

 

 

Sasatomi

124

12

71

1

10

1122

0

6

1346

 

 

Wu

84

8

43

1

6

865

1

0

1008

 

 

Totals

 

 

 

 

 

 

6986

 

 

 

 

 

 

 

 

 

 

 

 

 

 

The Division is involved in the following research projects and receives annual funding as follows:

 

Alloantibodies in Cardiac Transplantation Intervention, Outcomes and Mechanisms                           $131,779

Peripheral Mechanisms of Immunologic Tolerance                                                                                       $131,445

ITN Liver Analysis Core                                                                                                                                     $ 28,457

Hepatitis B Clinical Research Network Data Coordinating Center                                                               $   6,421

Targeting p27kip1 to Improve Hepatocyte Transplantation                                                                         $117,250

Luitpold Research Agreement                                                                                                                           $  42,599

B Cells in the Pathogenesis of Allograft Rejection                                                                                        $  14,264

Polyomavirus Infection after Kidney Transplantation                                                                                  $212,798

Immune Response to Polymavirus Infection                                                                                                   $240,178

In Vitro Screening of Drugs for Anti BK Viru Activity using Molecular Biology and

    Flow Cytometry Assays Applied to a Cell Culture System                                                                      $156,703

CPLA2, COX2 and PPAR-y in Cholangiocarcinoma                                                                                      $   40,042

Omega 3 Fatty Acids and Hepatic Carcinogenesis                                                                                        $   69,167

CPLA2A COX2 and TGF B in Liver Cancer                                                                                                     $212,500

Prostaglandin Signaling Pathway in Liver Cancer                                                                                          $  78,304

Alloantibodies in Cardiac Transplantation Intervention Outcomes and Mechanisms                            $136,584

Predictors for Drug Selection and Minimization in Pediatric liver Transplantation                            $ 9,366

Molecular Biology of Hemorrhagic Shock

   (Project V: Predictive Mathematical Model of Inflammation for Shock Trauma)                            $ 18,760

Core B Immunological Monitoring: Optimizing Outcome after Pediatric Heart Transplantation            $   67,687

Proj 3: Genetic Contributions to Graft and Patient Outcomes
Optimizing outcome after pediatric heart transplantation                                                                             $155,367

Total                                                                                                                                                                 $1,869,668

Research Activities

The Division has a strong commitment to research in transplantation.  Each professional is actively engaged in research projects frequently in collaboration with members of other departments, in particular, Transplant Surgery.  Over the year, faculty of the Division contributed 48 refereed publications, invited reviews and book chapters.  Forty-five invited lectures and scientific meeting presentations were given outside the institution, including 20 talks abroad.  Several members serve on grant review committees and editorial boards of major professional journals.  The division also operates a research histology laboratory for the Thomas E. Starzl Transplant Institute.

 

Current research activities deal with a variety of projects:

 

1.        Studies on the role of IL-6 in liver disease have shown that this cytokine is essential for maintaining biliary tree integrity and wound repair under conditions of biomedical stress, such as obstructive cholangiopathy.  Using IL-6 deficient mice, a model of accelerated development and decompensation of biliary cirrhosis has been established.  Analysis of IL-6/gp/30-dependent transcription factor activation is being used to determine how these signaling pathways contribute to the phenotype observed.  We are currently focusing on the functional characterization of an IL-6/STAT3-dependent family of molecules, small proline rich proteins, which function as SH3 domain ligands that protect cells from oxidative stress and induce epithelial-mesenchymal transition in biliary epithelial cells.

2.        We are currently evaluating tissue specimens from operationally tolerant liver allograft recipients that have been off of all immunosuppression for a period of 2.5 to 18 years after liver transplantation.  Multiplex staining modalities are being used to characterize the functional status of endothelial, parenchymal, and leukocyte populations within the tolerated organs to determine if, and how, the allograft contributes to allograft acceptance.

3.        Alloantibodies in Cardiac Transplantation – Intervention, Outcomes and Mechanisms Antibodies directed against allograft antigens are being increasingly recognized as critical determinants of allograft outcomes in adults. Pediatric heart transplant candidates are frequently allosensitized based on prior exposure to blood products and homografts. To overcome the high wait-list mortality, transplantation across a donor-specific cross-match has recently been performed in several pediatric centers, with early encouraging results. This application brings together a group of six leading heart transplant centers and leading transplantation scientists to study the impact of preformed and de novo alloantibodies on pediatric heart transplant outcomes.

4.        The Research Histology Laboratory of the Division serves as the Core Laboratory for two separate PO1 grants and one NIH contract:  1) Liver Tissue Core Laboratory Contract for the Immune Tolerance Network (ITN); 2) Histopathology Core for the PO1 grant entitled, “Peripheral Mechanisms of Toleranceâ€�, Dan Rothstein, PI; and 3) the Histopathology Core the PO1 grant entitled, “Allo-antibodies in Cardiac Transplantation-Intervention, Outcomes and Mechanismsâ€�.   Specific Web-based software has been developed to communicate results to investigators throughout the world on a real time basis.  Data management tools are being developed for this application.

5.        The division also maintains a robust clinical database, as a part of EDIT (Electronic Data Interface for Transplantation).  This database links histology diagnoses and histologic finding with a multitude of clinical, serologic, matching, donor and outcome parameters.  The database is an invaluable resource for clinical and patient-based research activities.

6.        Dendritic cells play an important role in inflammatory diseases, including allograft rejection, and steatohepatitis, as well as in tolerance to solid organ allografts.  IL-6/gp130 signaling plays an important role in their maturation.  Continuing studies are investigating the role of IL-6 dependent co-factors in dendritic cells maturation.

7.        Serological studies deal with the characterization of antibodies against private and public HLA epitopes and their correlation with amino acid residues.  Screening of sera from highly sensitized patients permits the identification of potential donors with acceptable mismatches and the assessment of the relative immunogenicity HLA antigens.

8.        A computer algorithm has been developed (HLA Matchmaker) that assesses donor-recipient HLA compatibility at the amino acid structural level.  The algorithm makes intralocus and interlocus comparisons of amino acid triplets of HLA class I antigens.

9.        Lung Transplantation is the only therapeutic option for many fatal lung diseases.  The current immunosuppression protocols based on systemic cyclosporine or tacrolimus are not adequate to control rejection.  A unique option is to target a high concentration of the immunosuppressive drugs to the lung allograft without increasing the systemic levels that might be toxic.  Studies are conducted in lung transplant patients that receive aerosol CsA and in control patients on the role of various cytokines and effector molecules that can cause allograft damage and dysfunction.  We also evaluate the impact of aerosol CsA on suppressing alloreactive T cells, on the alloantibody formation and release of soluble donor derived HLA antigen.  We are correlating the laboratory finding with the clinical parameters and determining whether these tests can be used as early markers of allograft damage.

10.     Post-transplant lymphoproliferative disease cytokine analysis of autologous EBV infected B cells has indicated Th2 type responsiveness during PTLD and Th1 type responses in normal EBV seropositive individuals. 

11.     Autologous lymphokine-activated killer (LAK) cell therapy has been successfully used to treat lymphoproliferative disorders arising in organ transplant patients.

12.     The clinical profile of patients with BK virus infection in the renal allograft is defined.  DNA sequencing of viral strains is being used to define mutations and genomic rearrangements, which may be relevant to the pathogenesis of an extremely refractory interstitial nephritis seen in these individuals.  Quantitative PCR assays have been developed to measure viral load in biopsy samples and body fluids to better manage the immunosuppression levels in these patients.  The interplay between host and viral immune factors in this disease is being investigated by DNA microarray technology.  Drugs with antiviral activity are being screened in an in-vitro culture model for possible future use in the clinical arena.

13.     Studies on human hepatocytes have shown that IL-6 and HGF regulate the activity of cytosolic phospholipase A2 (cPLA2) in cultured human hepatocytes.  The regulation of cPLA2 by cytokines and growth factors may play a role in the pathophysiology of inflammatory and non-inflammatory liver diseases.

14.     Human hepatobiliary cancers show increased expression of cyclooxygenase-2 (COX-2).  Studies have demonstrated that activation of Akt is an important mechanism by which COX-2 promotes hepatobiliary cancer growth.  Interruption of the COX-2 and prostaglandin signaling pathways is being targeted for the chemoprevention and treatment of human hepatobiliary cancers. 

15.     Peroxisome proliferator activated receptor-gamma (PPAR-gamma) ligands have been shown to inhibit human cholangiocarcinoma growth via a p53-dependent mechanism.  These pharmacological agents may represent promising therapeutic agents for the treatment of human cholangiocarcinoma.   

16.     Targeting p27kip1 to improve hepatocyte transplantation research.  In characterizing the cell-cycle regulatory role of the cyclin-dependent kinase inhibitor, p27kip1, it was discovered that p27 knockout mice exhibit pronounced multi-organomegaly with no increased spontaneous carcinogenesis in the liver.

17.     Optimizing outcome after pediatric heart transplantation.  This project focuses on different aspects of pediatric heart transplantation and how to optimize outcomes and minimize drug-related complications.

18.     We are conducting a multi-center project in pediatric heart transplant recipients to assess the impact of polymorphism in candidate genes that alter the frequency and severity of allograft rejection and the impact of pharmacogenomic factors that influence the drug efficacy, toxicity and drug dosing. The genetic polymorphism of pro-inflammatory cytokines and mediators as well as of growth factors is also of interest in lung transplantation.

19.     Another important area of research is the pre- and post-transplant sensitization of lung and other solid organ transplant recipients. We are interested to evaluate the development of anti-HLA and non-HLA antibodies in transplant recipients and the impact of various immunosuppressive strategies. The overall immuno-competence and the specific memory responses to various pathogens is an important aspect of patient management post-transplantation and assays are developed to assess the risk of infection and the extend for prophylaxis required based on the immune reactivity.

20.     Clinical research of antibody-mediated immune response in solid-organ transplantation is another area of investigation. The detection and specificity analysis of pre-formed and post-transplant developed antibodies are important components of humoral allo-response dynamics, while outcome association reveals the clinical impact of humoral/mixed forms of graft rejection. I am also interested in additional markers of antibody-mediated rejection (like soluble forms or vascular C4d deposition), as well as in the influence of various therapeutic protocols on HLA and non-HLA antibody response, both in adult and pediatric transplant populations.

21.     Another area of interest consists in the histocompatibility degree and its influence on allograft outcome. Together with Dr Duquesnoy, we are analyzing the HLA amino-acid residue matching using the HLAMatchmaker program, and we seek the impact on humoral sensitization. We are also using this algorithm for donor search in bone marrow and cord blood unit hematopoetic stem cell transplantation, in graft-versus-host and host-versus-graft disease, as well as for engraftment/chimerism studies and for virtual cross-matching in highly sensitized transplant candidates.

22.     Role of Bid protein on hepatic carcinogenesis in HBsAg transgenic mice.  Collaborative work with Xiao-Ming Yin, MD, PhD Associate Professor of Pathology, Division of Molecular Diagnostics, Department of Pathology, University of Pittsburgh School of Medicine.

23.     Assessment of transplant vasculopathy in mice that overexpress eNOS in the vasculature. Collaborative work with Timothy R. Billiar, MD, George Vance Foster Professor and Chair, Department of Surgery, University of Pittsburgh Medical Center.

24.     Participation in the Hepatitis B Research Network (http://www.hepbnet.org/) as pathologist member of  Data Coordinating Committee (Steve Belle Ph.D., P.I. RFA-DK-07-011) and member of Pathology Subcommittee. The network comprises a long-term multicenter study of hepatitis B patients to further clarify the biology and clinical progression of this disease, and carry out a number of clinical trials with supportive scientific studies to examine the efficacies of newer antiviral drugs and drug combinations.

Teaching/Training Activities

The Division is strongly devoted to teaching and training in transplantation pathology.  Professional staff directed graduate students and medical students in subjects, such as Cancer Biology, Digestion & Nutrition, Immunology & Inflammation, Kidney Pathology, and Molecular Pathology.  Visitors, including fellows and pathologists from various geographic areas of the United States, New Zealand, Germany and Canada to the Division for further training in transplant pathology.  Currently, the Tissue Typing Laboratory, under the direction of Adriana Zeevi, PhD. is training Dr. John Lunz as a post-doctoral fellow in Tissue Typing and Transplant Immunology.

 

The division also heads the Banff Working Group on Liver Allograft Pathology.  This group sets international guidelines for liver allograft biopsy interpretation and meets every two years to present new findings in the field and discuss future directions.

 

The Division is represented on the Diseases Transmission Advisory Committee of the OPTN/UNOS which reviews and provides guidance for cases of suspected donor disease transmission to transplant recipients. The committee is presently assembling a resource document to serve as a central information source regarding transplantation in this circumstance.

 

Many pathology residents have rotated through the Division to receive training in transplantation, hepatic and renal pathology.  This rotation is designed to provide a basic understanding of the molecular, cellular and histopathologic mechanisms of transplant immunity, allograft rejection opportunistic infections, organ preservation injury, recurrent disease, drug toxicity graft versus host disease and lymphoproliferative disease.  During this rotation, the residents attended various transplant conferences, research meetings and special lectures.

 

The Division conducts weekly interdisciplinary conferences on heart, kidney, kidney/pancreas, intestine, liver and native liver pathology.  Conferences in conjunction with Transplant Surgery deal with liver transplant tumors and Transplant Grand Rounds.  There are also bimonthly meetings to discuss research progress within the Division and weekly journal clubs.

 

Transplant Pathology Internet Service (TPIS)

 

Transplant Pathology Internet Services (TPIS): An Interactive Platform for the Standardization of Transplantation Pathology Practice, is a freely available worldwide web-based venture of the Division of Transplantation Pathology and was originally designed to foster collaboration and diagnostic standardization among transplant institutions. The entire site has been re-written using JAVA .jsp technology to replace the original combination of static html with occasional .cgi scripting. This has allowed more dynamic content, including the viewing of Whole Slide digital image photomicroscopy with zoom and annotation capability, as well as a richer and more responsive user experience.

This upgrade has also increased maintainability and enhancement possibilities to allow us to better fulfill the site's educational purpose.

 

TPIS presently comprises 1,986 pages/documents and has evolved into an educational and interactive medium promoting both communication and standardization of transplant pathology practice. It performs this role using a three-pronged approach:

 

  • The first and largest component of TPIS is didactic and follows classical web-based design. Thirty-two separate transplant-related histologic grading systems are incorporated with hyperlinked photomicrographs, serving as a de facto standard for biopsy interpretation. One hundred sixty-five separate consult cases, stripped of patient identifiers, are presented with full discussion. In addition, 8 separate didactic sections covering the handling and pathology of all major transplant organs are presented. A full-length textbook, Diagnostic Liver Pathology by Randall Lee is incorporated into the website.  An immunopathology module provides complementary information for the transplant pathologist. Several video lectures are also available on the site.

 

  • The second component comprises interactive education. A case conference module containing over 100 separate transplant-related cases represents a major portion of this section. New cases employ the use of whole slide digital imaging in place of selected .jpg images. (A separate electronic forum also serves an educational purpose, but has found its main use as a collaborative tool). A separate histopathologic testing module that can be modified for self-assessment or centralized scoring was added previously to conduct pathologist standardization testing for an international clinical drug trial. This approach, which can also be used for continuing medical education, has applicability for multi-center studies involving pathologists.

 

·         The third component that encourages standardized practice is an interactive portion directed toward altering the practice of the individual pathologist. In the simplest case, TPIS serves as a download site for specialized software. We have hosted software that performs HLA matching using the CREG approach and written within our group, and we are also offering telepathology software on a contractual basis. A second approach uses Java applets to perform medical calculations, allowing the user to perform calculations directly through the web browser. We have integrated five separate formulas that calculate creatinine clearance based on different clinical variables and offer all of these within our site. Similar calculations can be performed for determination of hepatic iron index.

 

 

For the one year period from July 1 2008 to June 30 2009, TPIS has received 143,592 page views from 132 different countries. (The top 10 countries in order are: US, Greece, Italy, Ireland, Canada, Germany, United Kingdom, India, Czech Republic, and Japan). The average visitor views 4.1 pages per visit and spends an average of 3 minutes and 10 seconds per visit. There have been a total of 35,177 separate visits with 54% representing new, and 46% representing returning, visitors. The importance of this site is highlighted by the fact that 3,484 unique users have accessed TPIS more than 200 times during this interval, with another 704 using it between 1-200 times, and 718 between 51-100 times. Traffic derives from direct URL access in 83% of cases, with another 9.9% coming from search engines and 7.7% from referring sites. The top 5 search terms were: electron dense deposits, transplant pathology, HAI score, NAFLD activity score, and rejection activity index. The top referral sites included: images.google.com, pathresident.dhe.duke.edu, pulse.uphs.upenn.edu, and pathguy.com. TPIS itself also contains an integrated search engine that allows a shortcut to specific content. The top 10 search terms are: GVHD, Liver, creatinine clearance, C4d, CMV, kidney, Banff, PTLD, steatohepatitis, and hepatitis.

When searching on Google for 'Transplant Pathology' - TPIS is the first returned link, showing strong usage and correlation for its intended audience.