Online pathology request platform at the Semmelweis University / Online patológiai vizsgálatkéro felület létrehozása a Semmelweis Egyetemen.

Összefoglaló. Bevezetés: Az I. Patológiai és Kísérleti Rákkutató Intézet - a Semmelweis Egyetemen belüli diagnosztikai szolgáltatásnyújtás mellett - kiterjedt külso partneri hálózattal (vizsgálatmegrendelovel) bír. Az Intézet a napi muködése során párhuzamosan használja az egyetem központi informatikai rendszerét, valamint belso, folyamattámogató alkalmazását (workflow management). A külsos partnerek hozzáférése vizsgálatfeladásra az egyetemi központi informatikai rendszerhez nincs biztosítva. A vizsgálatok rendelése papíralapú, a minta érkeztetésekor a klinikai adatok rögzítése manuális, kifejezetten humáneroforrás-igényes. Célkituzés: Célunk volt a patológiai minták regisztrációjának egyszerusítése és felgyorsítása, az adminisztratív folyamatok hatékonyságának javítása. Módszer: A kituzött célt a minoségfejlesztésbol ismert Plan-Do-Check-Act (Tervezés-Cselekvés-Ellenorzés-Beavatkozás) ciklus módszereit alkalmazva kívántuk elérni, online, a mintavétel helyén elérheto, a meglévo belso folyamattámogató alkalmazáshoz kapcsolódó, szakterület-specifikus vizsgálatkéro felület kifejlesztésével. Eredmények: A vizsgálati minták regisztrációjának átlagos ideje 65%-kal csökkent az online vizsgálatkéro rendszerhez csatlakozott klinikai partnerek körében. Megbeszélés: Az elmúlt években tapasztalható volt, hogy kisebb, nem hatékonyan muködtetheto patológiai osztályok megszuntek, részben vagy egészben beolvadtak nagyobb diagnosztikai egységekbe. A humáneroforrás-problémák (elöregedo szakma a patológia) a fenti folyamatot minden bizonnyal tovább erosítik. Várható, hogy a nagyobb patológiai osztályokon a következo években a mintaszám tovább növekszik, a vizsgálatkérések egyre nagyobb hányada érkezik majd intézményen kívülrol. Következtetés: A patológiai informatika fejlesztésekor figyelembe kell venni, hogy szükséges már a mintavétel helyén biztosítani az informatikai támogatást a minta nyomon követéséhez, nem elégséges csak a laboron belüli folyamatok kiszolgálása. Orv Hetil. 2021; 162(49): 1962-1967. INTRODUCTION: The 1st Department of Pathology and Experimental Cancer Research, Semmelweis University (Budapest, Hungary) has a broad network of clinical partners, many of which are non-university hospitals. A separate hospital information system and a local laboratory workflow management system is used at the Department. University clinics use the hospital information system for electronic requesting of tests. Non-university partners have no access to the systems, requesting tests is paper-based, registration of the requests at the pathology lab is manual and laborious. OBJECTIVE: Our main objective was to improve the efficiency of the sample registration step of the pathology workflow. METHOD: Applying the Plan-Do-Check-Act procedure, a quality improvement project has been carried out and an online, subspecialty-based requesting application tool, interfaced with the current laboratory information system, was developed. RESULTS: The average sample registration time improved with 65% among the early user partners. DISCUSSION: The past years have shown smaller, inefficient pathology labs decreasing in number and integrated into larger regional diagnostic centers. Both issues of efficiency and quality assurance and problems rooted in human resources are drivers of further centralisation. The numbers of test requests and samples from non in-house partners are expected to be increased in the pathology labs in the future. CONCLUSION: Efficient and safe sample tracking has to start at the site of sample acquisition. State of the art laboratory information systems should support this expansion of competence. Orv Hetil. 2021; 162(49): 1962-1967.

Diagnostic problems of the mucormycosis of the maxillary sinus Case presentation.

Mucormycosis is a fulminant opportunistic infection with significant mortality in susceptible individuals. Although mortality rates vary widely (30 to 100%) according to professional literature, recently in instances with no central nervous system (CNS) involvement the survival rate averages varies between 50 and 80% owing to complex therapy. With CNS involvement, however, the fatality rate is over 80%. Predisposing diseases include diabetes mellitus, malnutrition, hematologi- cal diseases, neutropenia, burns, surgical procedures, antibiotic treatments, long-term steroid therapy and immunosuppressive therapy. Mucormycosis may at times arise even in -immunocompetent individuals. It has diverse clinical forms with the most frequent forms being rhino-maxillary and rhino-oculocerebral (the latter of which is characterized by a high mortality rate). They mainly enter the body through inhalation, with saprophytic mucor species often demonstrable in the upper respiratory tracts, which are nevertheless non-pathogenic in most healthy individuals. The spores may also enter percutaneously through traumas, skin lesions, insect bites, or injections (e.g. through intravenous drug use); as well as via the alimentary tract with contaminated foodstuff. The prognosis can be improved by a quick establishment of the diagnosis, the quick initiation of the therapy and treatment of the underlying disease. Although first and foremost the recognition and treatment of the disease does not rest with dentists and oral surgeons, in order to localize the disease it is important to examine it from a differential diagnostic point of view and interdisciplinary cooperation may also be required in a complex treatment. Our aim is to introduce mucormycosis in our case report study.

[First attempts in the introduction of cryopreservation of ovarian tissues]. / Kezdeti tapasztalataink a petefészekszövet-fagyasztás bevezetésével.

INTRODUCTION: The oncological treatment may damage ovarian function. To prevent this, it is possible to cryopreserve the ovarian tissue, and to keep the samples for long-term storage. The frozen-thawed tissue could be retransplanted after chemo- or radiotherapy. AIM: The aim of our study was to examine the effect of cryopreservation on the viability of ovarian tissue. METHOD: We analyzed the survival of frozen-thawed donated ovarian tissues. The quality of the follicles and hormone production in fresh and frozen-thawed samples were compared. RESULTS: Histological analysis showed that the number of viable follicles was reduced by 23% in the frozen-thawed samples. However, viable follicles still presented in post thawing ovarian tissues. Maximal estradiol production in frozen-thawed tissues was 908 pg/ml and hormone production was similar to the control tissues. The maximal progesterone production was 1.95 ng/ml post thawing, but these values were lower than the progesterone production of fresh tissues. CONCLUSIONS: The method of ovarian cryopreservation used in our laboratory was able preserve the viability of follicles in frozen-thawed ovarian tissues. Orv. Hetil., 2016, 157(49), 1947-1954.

3-dimensional digital reconstruction of the murine coronary system for the evaluation of chronic allograft vasculopathy.

BACKGROUND: Chronic allograft vasculopathy (CAV) is a major mechanism of graft failure of transplanted organs in humans. Morphometric analysis of coronary arteries enables the quantitation of CAV in mouse models of heart transplantation. However, conventional histological procedures using single 2-dimensional sections limit the accuracy of CAV quantification. The aim of this study is to improve the accuracy of CAV quantification by reconstructing the murine coronary system in 3-dimensions (3D) and using virtual reconstruction and volumetric analysis to precisely assess neointimal thickness. METHODS: Mouse tissue samples, native heart and transplanted hearts with chronic allograft vasculopathy, were collected and analyzed. Paraffin embedded samples were serially sectioned, stained and digitized using whole slide digital imaging techniques under normal and ultraviolet lighting. Sophisticated software tools were used to generate and manipulate 3D reconstructions of the major coronary arteries and branches. RESULTS: The 3D reconstruction provides not only accurate measurements but also exact volumetric data of vascular lesions. This virtual coronary arteriography demonstrates that the vasculopathy lesions in this model are localized to the proximal coronary segments. In addition, virtual rotation and volumetric analysis enabled more precise measurements of CAV than single, randomly oriented histologic sections, and offer an improved readout for this important experimental model. CONCLUSIONS: We believe 3D reconstruction of 2D histological slides will provide new insights into pathological mechanisms in which structural abnormalities play a role in the development of a disease. The techniques we describe are applicable to the analysis of arteries, veins, bronchioles and similar sized structures in a variety of tissue types and disease model systems. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/3772457541477230 .

Poorly differentiated synovial sarcoma is associated with high expression of enhancer of zeste homologue 2 (EZH2).

BACKGROUND: Enhancer of zeste homologue 2 (EZH2) is a polycomb group (PcG) family protein. Acting as a histone methyltransferase it plays crucial roles in maintaining epigenetic stem cell signature, while its deregulation leads to tumor development. EZH2 overexpression is commonly associated with poor prognosis in a variety of tumor types including carcinomas, lymphomas and soft tissue sarcomas. However, although the synovial sarcoma fusion proteins SYT-SSX1/2/4 are known to interact with PcG members, the diagnostic and prognostic significance of EZH2 expression in synovial sarcoma has not yet been investigated. Also, literature data are equivocal on the correlation between EZH2 expression and the abundance of trimethylated histone 3 lysine 27 (H3K27me3) motifs in tumors. METHODS: Immunohistochemical stains of EZH2, H3K27me3, and Ki-67 were performed on tissue microarrays containing cores from 6 poorly differentiated, 39 monophasic and 10 biphasic synovial sarcomas, and evaluated by pre-established scoring criteria. Results of the three immunostainings were compared, and differences were sought between the histological subtypes as well as patient groups defined by gender, age, tumor location, the presence of distant metastasis, and the type of fusion gene. The relationship between EZH2 expression and survival was plotted on a Kaplan-Meier curve. RESULTS: High expression of EZH2 mRNA and protein was specifically detected in the poorly differentiated subtype. EZH2 scores were found to correlate with those of Ki-67 and H3K27me3. Cases with high EZH2 score were characterized by larger tumor size (≥ 5cm), distant metastasis, and poor prognosis. Even in the monophasic and biphasic subtypes, higher expression of EZH2 was associated with higher proliferation rate, larger tumor size, and the risk of developing distant metastasis. In these histological groups, EZH2 was superior to Ki-67 in predicting metastatic disease. CONCLUSIONS: High expression of EZH2 helps to distinguish poorly differentiated synovial sarcoma from the monophasic and biphasic subtypes, and it is associated with unfavorable clinical outcome. Importantly, high EZH2 expression is predictive of developing distant metastasis even in the better-differentiated subtypes. EZH2 overexpression in synovial sarcoma is correlated with high H3K27 trimethylation. Thus, along with other epigenetic regulators, EZH2 may be a future therapeutic target.

A challenging case of metastatic intra-abdominal synovial sarcoma with unusual immunophenotype and its differential diagnosis.

The primary and metastatic gastrointestinal synovial sarcoma is rare with a wide differential diagnosis. It usually expresses cytokeratins EMA, BCL2 with an occasional CD99, and S100 positivity but not desmin. We present a case of metastatic synovial sarcoma with unusual immunophenotype causing diagnostic challenges. The tumor cells showed focal cytokeratin, EMA, and, unexpectedly, desmin positivity. Additional intranuclear TLE-1 positivity and negativity for CD34 and DOG-1 were also identified. A diagnosis of monophasic synovial sarcoma was confirmed by using FISH break-apart probe. RT-PCR revealed the SYT-SSX1 fusion gene. Intra-abdominal synovial sarcoma, either primary or metastatic, with unusual desmin positivity raises the diagnostic challenge, since a wide range of differential diagnoses could show a similar immunophenotype (leiomyosarcoma, desmoid tumor, myofibroblastic tumor, and rarely GIST etc.). Typical morphology and focal cytokeratin/EMA positivity should alert to this tumor, and FISH and RT-PCR remain the gold standard for the confirmation.

Gonadoblastoma: Case report of two young patients with isochromosome 12p found in the dysgerminoma overgrowth component in one case.

Gonadoblastomas are unusual neoplasias that frequently appear in the dysgenetic gonads of women with chromosome Y anomaly. We present two cases of gonadoblastoma associated with complete gonadal dysgenesis and Turner syndrome, respectively, with dysgerminoma overgrowth found in one case. We were interested in the DNA ploidy, the presence of Y chromosome DNA sequence and the status of chromosome 12p arm among the tumor cells. We performed cytophotometry to analyze the DNA content and fluorescence in situ hybridization (FISH) to identify the Y chromosome and the isochromosome 12p within the tumor cells. The cytophotometric result showed diploid DNA content in gonadoblastoma, whereas dysgerminoma revealed aneuploid DNA. The FISH result revealed Y chromosome DNA sequence within gonadoblastoma and dysgerminoma. Isochromosome 12p was identified in dysgerminoma, but not in gonadoblastoma. We conclude that gonadoblastoma and dysgerminoma have a strong association with the Y chromosome, and dysgerminoma overgrowth is due to further chromosomal aberrations, such as isochromosome 12p. Histological, immunohistocheimcal and molecular studies should render the correct diagnosis. Identifying dysgerminoma overgrowth is crucial since it is associated with adverse prognosis and requires additional therapy.

Validation of diagnostic accuracy using digital slides in routine histopathology.

BACKGROUND: Robust hardware and software tools have been developed in digital microscopy during the past years for pathologists. Reports have been advocated the reliability of digital slides in routine diagnostics. We have designed a retrospective, comparative study to evaluate the scanning properties and digital slide based diagnostic accuracy. METHODS: 8 pathologists reevaluated 306 randomly selected cases from our archives. The slides were scanned with a 20× Plan-Apochromat objective, using a 3-chip Hitachi camera, resulting 0.465 µm/pixel resolution. Slide management was supported with dedicated Data Base and Viewer software tools. Pathologists used their office PCs for evaluation and reached the digital slides via intranet connection. The diagnostic coherency and uncertainty related to digital slides and scanning quality were analyzed. RESULTS: Good to excellent image quality of slides was recorded in 96%. In half of the critical 61 digital slides, poor image quality was related to section folds or floatings. In 88.2% of the studied cases the digital diagnoses were in full agreement with the consensus. Out of the overall 36 incoherent cases, 7 (2.3%) were graded relevant without any recorded uncertainty by the pathologist. Excluding the non-field specific cases from each pathologist's record this ratio was 1.76% of all cases. CONCLUSIONS: Our results revealed that: 1) digital slide based histopathological diagnoses can be highly coherent with those using optical microscopy; 2) the competency of pathologists is a factor more important than the quality of digital slide; 3) poor digital slide quality do not endanger patient safety as these errors are recognizable by the pathologist and further actions for correction could be taken. VIRTUAL SLIDES: The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/1913324336747310.

Correlation between DNA ploidy, metaphase high-resolution comparative genomic hybridization results and clinical outcome of synovial sarcoma.

BACKGROUND: Although synovial sarcoma is the 3rd most commonly occurring mesenchymal tumor in young adults, usually with a highly aggressive clinical course; remarkable differences can be seen regarding the clinical outcome. According to comparative genomic hybridization (CGH) data published in the literature, the simple and complex karyotypes show a correlation between the prognosis and clinical outcome. In addition, the connection between DNA ploidy and clinical course is controversial. The aim of this study was using a fine-tuning interpretation of our DNA ploidy results and to compare these with metaphase high-resolution CGH (HR-CGH) results. METHODS: DNA ploidy was determined on Feulgen-stained smears in 56 synovial sarcoma cases by image cytometry; follow up was available in 46 cases (average: 78 months). In 9 cases HR-CGH analysis was also available. RESULTS: 10 cases were found DNA-aneuploid, 46 were DNA-diploid by image cytometry. With fine-tuning of the diploid cases according to the 5c exceeding events (single cell aneuploidy), 33 cases were so called "simple-diploid" (without 5c exceeding events) and 13 cases were "complex-diploid"; containing 5c exceeding events (any number). Aneuploid tumors contained large numbers of genetic alterations with the sum gain of at least 2 chromosomes (A-, B- or C-group) detected by HR-CGH. In the "simple-diploid" cases no or few genetic alterations could be detected, whereas the "complex-diploid" samples numerous aberrations (equal or more than 3) could be found. CONCLUSIONS: Our results show a correlation between the DNA-ploidy, a fine-tuned DNA-ploidy and the HR-CGH results. Furthermore, we found significant correlation between the different ploidy groups and the clinical outcome (p < 0.05).

Shifting gears higher--digital slides in graduate education--4 years experience at Semmelweis University.

BACKGROUND: The spreading of whole slide imaging or digital slide systems in pathology as an innovative technique seems to be unstoppable. Successful introduction of digital slides in education has played a crucial role to reach this level of acceptance. Practically speaking there is no university institute where digital materials are not built into pathology education. At the 1st. Department of Pathology and Experimental Cancer Research, Semmelweis University optical microscopes have been replaced and for four years only digital slides have been used in education. The aim of this paper is to summarize our experiences gathered with the installation of a fully digitized histology lab for graduate education. METHODS: We have installed a digital histology lab with 40 PCs, two slide servers - one for internal use and one with external internet access. We have digitized hundreds of slides and after 4 years we use a set of 126 slides during the pathology course. A Student satisfaction questionnaire and a Tutor satisfaction questionnaire have been designed, both to be completed voluntarily to have feed back from the users. The page load statistics of the external slide server were evaluated. RESULTS: The digital histology lab served ~900 students and ~1600 hours of histology practice. The questionnaires revealed high satisfaction with digital slides. The results also emphasize the importance of the tutors' attitude towards digital microscopy as a factor influencing the students' satisfaction. The constantly growing number of page downloads from the external server confirms this satisfaction and the acceptance of digital slides. CONCLUSIONS: We are confident, and have showed as well, that digital slides have got numerous advantages over optical slides and are more suitable in education.

Strong synergy of heat and modulated electromagnetic field in tumor cell killing.

BACKGROUND AND PURPOSE: Hyperthermia is an emerging complementary method in radiooncology. Despite many positive studies and comprehensive reviews, the method is not widely accepted as a combination to radiotherapy. Modulated electrohyperthermia (mEHT; capacitive, electric field modulated, 13.56 MHz) has been used in clinical practice for almost 2 decades in Germany, Austria and Hungary. This in vivo study in nude mice xenograft tumors compares mEHT with "classic" radiative hyperthermia (radHT). MATERIAL AND METHODS: Nude mice were xenografted with HT29 human colorectal carcinoma cells. 28 mice in four groups with seven animals each and two tumors per animal (totally 56 tumors) were included in the present study: group 1 as untreated control; group 2 treated with radHT at 42 degrees C; group 3 treated with mEHT at identical 42 degrees C; group 4 treated with mEHT at 38 degrees C (by intensively cooling down the tumor). 24 h after treatment, animals were sacrificed and the tumor cross sections studied by precise morphological methods for the respective relative amount of "dead" tumor cells. RESULTS: The effect of mEHT established a double effect as a synergy between the purely thermal (temperature-dependent) and nonthermal (not directly temperature-dependent) effects. The solely thermal enhancement ratio (TER) of cell killing was shown to be 2.9. The field enhancement ratio (FER) at a constant temperature of 42 degrees C was measured as 3.2. Their complex application significantly increased the therapeutic enhancement to 9.4. CONCLUSION: mEHT had a remarkable cancer cell-killing effect in a nude mice xenograft model.