Modifications of DAMPs levels in extracellular environment induced by aminolevulinic acid-based photodynamic therapy of esophageal cancer cells.

PURPOSE: Immunogenic cell death plays an important role in anticancer treatment because it combines cell death with appearance of damage associated molecular patterns that have the potential to activate anticancer immunity. Effects of damage associated molecular patterns induced by aminolevulinic acid-based photodynamic therapy were studied mainly on dendritic cells. They have not been deeply studied on macrophages that constitute the essential component of the tumor microenvironment. The aim of this study was to analyze features of esophageal cancer cell death in relation to release capacity of damage associated molecular pattern species, and to test the effect of related extracellular environmental alterations on macrophages. MATERIAL AND METHODS: Esophageal Kyse 450 carcinoma cells were subjected to aminolevulinic acid-based photodynamic therapy at different concentrations of aminolevulinic acid. Resting, IFN/LPS and IL-4 macrophage subtypes were prepared from monocytic THP-1 cell line. Cell death features and macrophage modifications were analyzed by fluorescence-based live cell imaging. ATP and HMGB1 levels in cell culture media were determined by ELISA assays. The presence of lipid peroxidation products in culture media was assessed by spectrophotometric detection of thiobarbituric acid reactive substances. RESULTS: Aminolevulinic acid-based photodynamic therapy induced various death pathways in Kyse 450 cells that included features of apoptosis, necrosis and ferroptosis. ATP amounts in extracellular environment of treated Kyse 450 cells increased with increasing aminolevulinic acid concentration. Levels of HMGB1, detectable by ELISA assay in culture media, were decreased after the treatment. Aminolevulinic acid-based photodynamic therapy induced lipid peroxidation of cellular structures and increased levels of extracellular lipid peroxidation products. Incubation of resting and IL-4 macrophages in conditioned medium from Kyse 450 cells treated by aminolevulinic acid-based photodynamic therapy induced morphological changes in macrophages, however, comparable alterations were induced also by conditioned medium from untreated cancer cells. CONCLUSION: Aminolevulinic acid-based photodynamic therapy leads to alterations in local extracellular levels of damage associated molecular patterns, however, comprehensive studies are needed to find whether they can be responsible for macrophage phenotype modifications.

Long COVID Complicated by Fatal Cytomegalovirus and Aspergillus Infection of the Lungs: An Autopsy Case Report.

After the acute phase of COVID-19, some patients develop long COVID. This term is used for a variety of conditions with a complex, yet not fully elucidated etiology, likely including the prolonged persistence of the virus in the organism and progression to lung fibrosis. We present a unique autopsy case of a patient with severe COVID-19 with prolonged viral persistence who developed interstitial lung fibrosis complicated by a fatal combination of cytomegalovirus and Aspergillus infection. SARS-CoV-2 virus was detected at autopsy in the lungs more than two months after the acute infection, although tests from the nasopharynx were negative. Immune dysregulation after COVID-19 and the administration of corticoid therapy created favorable conditions for the cytomegalovirus and Aspergillus infection that were uncovered at autopsy. These pathogens may represent a risk for opportunistic infections, complicating not only the acute coronavirus infection but also long COVID, as was documented in the presented case.

The Neglected Uterine NK Cells/Hamperl Cells/Endometrial Stromal Granular Cell, or K Cells: A Narrative Review from History through Histology and to Medical Education.

Reproductive immunology is at the forefront of research interests, aiming to better understand the mechanisms of immune regulation during gestation. The relationship between the immune system and the implanting embryo is profound because the embryo is semi-allogenic but not targeted by the maternal immune system, as expected in graft-versus-host reactions. The most prominent cell population at the maternal-fetal interface is the population of uterine natural killer (uNK) cells. Uterine NK cells are two-faced immunologically active cells, bearing comparison with Janus, the ancient Roman god of beginnings and endings. Their first face can be seen as natural killer cells, namely lymphocytes, which are critical for host defense against viruses and tumors. Even though uNK cells contain cytolytic molecules, their cytotoxic effect is not applied to classical target cells in vivo, playing a permissive rather than a defensive role. Their second face is crucial in maintaining physiological gestation-uNK cells show critical immunomodulatory functions with the potential to control embryo implantation and trophoblast invasion, regulate placental vascular remodeling, and promote embryonic/fetal growth. Therefore, we believe that their current designation "natural killer cells" (the first "cytotoxic" Janus's face) is misleading and inappropriate, considering their principal function is supporting and maintaining pregnancy. In this narrative review, we will focus on three lesser-known areas of knowledge about uNK cells. First, from the point of view of histology, we will comprehensively map the history of the discovery of these cells, as well as the current histological possibilities of their identification within the endometrium. To be brief, the discovery of uNK cells is generally attributed to Herwig Hamperl, one of the most influential and prominent representatives of German pathology in the 20th century, and his co-worker, Gisela Hellweg. Secondly, we will discuss the interesting aspect of terminology, since uNK cells are probably one of the human cells with the highest number of synonymous names, leading to significant discrepancies in their descriptions in scientific literature. From the first description of this cell type, they were referred to as endometrial granulocytes, granular endometrial stromal cells, or large granular lymphocytes until the end of the 1980s and the beginning of the 1990s of the last century, when the first publications appeared where the name "uterine NK cells" was used. The third area of present review is medical teaching of histology and clinical embryology. We can confirm that uNK cells are, in most textbooks, overlooked and almost forgotten cells despite their enormous importance. In the present narrative review, we summarize the lesser-known historical and terminological facts about uNK cells. We can state that within the textbooks of histology and embryology, this important cell population is still "overlooked and neglected" and is not given the same importance as in fields of clinical research and clinical practice.

Trichinella spiralis (Owen, 1835) Induces Increased Dystrophin Expression in Invaded Cross-striated Muscle.

PURPOSE: Dystrophin and the dystrophin glycoprotein complex serve as a cytoskeletal integrator, critical for muscle membrane stability. The aim of the present study was to clarify the expression of dystrophin protein and mRNA in the skeletal muscle tissue during the muscle phase of trichinellosis in mice. METHODS: Muscle tissue was collected from mice experimentally infected with Trichinella spiralis at days 0, 14 and 40 after infection. The expression of dystrophin in the muscle tissue was investigated by immunohistochemistry with antibodies against three different domains of the protein, and the expression levels of Dys mRNA by real-time PCR. RESULTS: The presence of dystrophin protein was increased in the de-differentiating cytoplasm at the early stage of muscle infection and was persisting also in the mature Nurse cell harbouring the parasite. It was accompanied by significantly elevated expression of Dys mRNA at days 14 and 40 after infection. CONCLUSION: Our findings indicate that dystrophin plays a role in regeneration of the muscle and in the Nurse cell formation and stability for security of the parasite survival.

Evaluation and limitations of different approaches among COVID-19 fatal cases using whole-exome sequencing data.

BACKGROUND: COVID-19 caused by the SARS-CoV-2 infection may result in various disease symptoms and severity, ranging from asymptomatic, through mildly symptomatic, up to very severe and even fatal cases. Although environmental, clinical, and social factors play important roles in both susceptibility to the SARS-CoV-2 infection and progress of COVID-19 disease, it is becoming evident that both pathogen and host genetic factors are important too. In this study, we report findings from whole-exome sequencing (WES) of 27 individuals who died due to COVID-19, especially focusing on frequencies of DNA variants in genes previously associated with the SARS-CoV-2 infection and the severity of COVID-19. RESULTS: We selected the risk DNA variants/alleles or target genes using four different approaches: 1) aggregated GWAS results from the GWAS Catalog; 2) selected publications from PubMed; 3) the aggregated results of the Host Genetics Initiative database; and 4) a commercial DNA variant annotation/interpretation tool providing its own knowledgebase. We divided these variants/genes into those reported to influence the susceptibility to the SARS-CoV-2 infection and those influencing the severity of COVID-19. Based on the above, we compared the frequencies of alleles found in the fatal COVID-19 cases to the frequencies identified in two population control datasets (non-Finnish European population from the gnomAD database and genomic frequencies specific for the Slovak population from our own database). When compared to both control population datasets, our analyses indicated a trend of higher frequencies of severe COVID-19 associated risk alleles among fatal COVID-19 cases. This trend reached statistical significance specifically when using the HGI-derived variant list. We also analysed other approaches to WES data evaluation, demonstrating its utility as well as limitations. CONCLUSIONS: Although our results proved the likely involvement of host genetic factors pointed out by previous studies looking into severity of COVID-19 disease, careful considerations of the molecular-testing strategies and the evaluated genomic positions may have a strong impact on the utility of genomic testing.

Suppression of resistance to aminolevulinic acid-based photodynamic therapy in esophageal cell lines by administration of iron chelators in collagen type I matrices.

PURPOSE: Photodynamic therapy (PDT) utilizes visible light to activate the cytotoxic effects of photosensitizing drugs. PDT protocols require optimization to overcome treatment resistance and induce a beneficial anti-tumor immune response. The aim of this study was to examine the possibility to suppress the resistance of esophageal cell lines to aminolevulinic acid (ALA)-PDT by administration of iron chelators to induce sufficient cell cytotoxicity under pathophysiologically relevant conditions, mimicking the advanced stages of cancer. MATERIALS AND METHODS: Effects of ALA-PDT in combination with iron chelators were compared in three esophageal cell lines in conventional monolayers and in 3 D cultures based on collagen type I. Modified colony assay and fluorescence-based live cell imaging, respectively were applied. The latter was used also to test the capability of pre-polarized macrophages to interact with cancer cells subjected to ALA-PDT with or without iron chelators. RESULTS: Iron chelators were effective in the enhancement of ALA-PDT in all cell lines under both culture conditions. Fluorescence evaluation of cell viability in 3 D cultures indicated the contribution of apoptotic cell death after ALA-PDT, both with and without iron chelators. Engulfment of remnants of dead cancer cells by macrophages in 2 D cultures was indicated, however, the interaction between macrophages and cancer cells in 3 D cultures subjected to ALA-PDT with or without iron chelators was not present. CONCLUSIONS: The potential of iron chelators to enhance ALA-PDT was maintained in 3 D collagen matrices. Although PDT dose (ALA concentration, light exposure time) required modification in a cell line-dependent manner to achieve a comparable effect of PDT alone in conventional monolayers and in collagen matrices, the potential of iron chelators to suppress the resistance of esophageal cells to ALA-PDT was not influenced by a fibrillar collagen matrix.

Innervation density and types of nerves in prostate cancer.

Innervation of cancerous tissue represents an important pathway enabling the nervous system to influence the processes associated with the initiation, progression, and metastasis of a neoplastic process. In the context of prostate cancer, several papers report the presence of innervation and its modulating effect on the cancer prognosis. However, most of the data are experimental, with limited information on human prostate cancer innervation. Morphometric analysis of archival prostate specimen immunohistochemistry with neural markers PGP9.5 and S100 showed a significant decrease of nerve density in the prostate cancer (n=44) compared to the normal prostate tissue (n=18) and benign prostatic hyperplasia (n=28). Sympathetic nerves were detected with TH, parasympathetic with VAChT, and sensory nerves with SP and CGRP protein detection. Dual immunofluorescence revealed numerous sympathetic nerves in normal prostate and benign prostatic hyperplasia, especially in the peripheral parts. Only a few parasympathetic nerves were found between the glands and in the peripheral parts of the prostate and benign hyperplasia. Sporadic positivity for sensory innervation was present only in approximately 1/10 of nerve fibers, especially in the larger nerves. The pattern of innervation in prostate cancer was analogous to that in normal prostate gland and benign prostatic hyperplasia but there was a significantly lower amount of all nerve types, especially in high-grade carcinoma cases. Although not significant, there was a tendency of decreasing innervation density with increasing Gleason score. Regarding the low density of nerves in prostate carcinoma, the significantly lower PCNA counts in nerves of the cancer specimens cannot be ascribed to lower proliferation activity. Our data confirmed the lower nerve density in the prostate cancer compared to the benign prostate tissue. We could not approve an increased nerve proliferation activity in prostate cancer. All nerve types, most the sympathetic, less the parasympathetic, and the sensory nerves, are present in prostate cancer. The highest nerve density at the periphery of the cancer tissue implies this to be the result of an expansive tumor growth. It is evident that the results of experimental prostate cancer models can be applied to human pathology only to a certain extent. The relation between the range of innervation and the biology of prostate cancer is very complex and will require more detailed information to be applied in therapeutic solutions.

Rare Location of a Dermoid Cyst in the Parotid Gland: A Case Report.

Dermoid cyst of the parotid gland is a lesion composed of benign tissues of ectodermal and mesodermal origin. Although a dermoid cyst can be encountered across nearly all sites of the body, its location in the head and neck area is quite uncommon and even more unusual inside the parotid gland. We present a case of a patient with gradually enlarging tumour in her right parotid gland who underwent surgical removal of the tumour histologically corresponding to a dermoid cyst.

Primary thyroid leiomyosarcoma with transvenous extension to the right atrium: a case report.

Background: Primary thyroid leiomyosarcoma (LMS) is a very rare tumour with less than 40 published cases yet. Direct metastatic extension into the great cervical veins and caval veins is extremely uncommon. Extension to the right heart has not yet been reported. Case summary: A 62-year-old man was admitted for sudden onset of left neck pain and dyspnoea. Computed tomography and ultrasonography found an extensive mass of the left great cervical veins, superior vena cava, and a nodule in the left thyroid lobe. Transesophageal echocardiography visualized large protrusion of this mass into the right atrium. Cytology of a thyroid nodule diagnosed a benign hyperplastic nodule. The mass was considered to be likely an extensive thrombus. The patient was started on anticoagulant therapy. The next course was complicated by pulmonary embolism and later by enterorrhagia. Despite clinical stabilization, the patient died suddenly. Autopsy finding differed from the clinical conclusion. Microscopic investigation revealed that the mass seen in the cervical veins down to the right atrium was a spindle cell tumour with a primary site in the left thyroid lobe. Immunohistochemistry was consistent with the final diagnosis of primary thyroid LMS. Discussion: Differential diagnosis of the masses of great cervical veins and right atrium can be challenging. Pure venous thrombus and tumour thrombus must be distinguished. Thyroid LMS should also be considered in patients with masses in the right atrium and thyroid nodules.

Ciliary Body Leiomyoma.

The report aims to present the case of intraocular leiomyoma. We conducted a case study on a patient who presented with an intraocular tumour. After examination, including magnetic resonance, positron emission tomography with computed tomography, B-scan, we performed surgery - enucleation of the eye globe with histological verification of tumour mass. Histological analysis of enucleated eyes proved intraocular leiomyoma. Leiomyoma is a rare intraocular tumour, which is clinically challenging to recognize; therefore, histological confirmation is most often required.

Placental Tissue Destruction and Insufficiency From COVID-19 Causes Stillbirth and Neonatal Death From Hypoxic-Ischemic Injury.

CONTEXT.­: Perinatal death is an increasingly important problem as the coronavirus disease 2019 (COVID-19) pandemic continues, but the mechanism of death has been unclear. OBJECTIVE.­: To evaluate the role of the placenta in causing stillbirth and neonatal death following maternal infection with COVID-19 and confirmed placental positivity for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). DESIGN.­: Case-based retrospective clinicopathologic analysis by a multinational group of 44 perinatal specialists from 12 countries of placental and autopsy pathology findings from 64 stillborns and 4 neonatal deaths having placentas testing positive for SARS-CoV-2 following delivery to mothers with COVID-19. RESULTS.­: Of the 3 findings constituting SARS-CoV-2 placentitis, all 68 placentas had increased fibrin deposition and villous trophoblast necrosis and 66 had chronic histiocytic intervillositis. Sixty-three placentas had massive perivillous fibrin deposition. Severe destructive placental disease from SARS-CoV-2 placentitis averaged 77.7% tissue involvement. Other findings included multiple intervillous thrombi (37%; 25 of 68) and chronic villitis (32%; 22 of 68). The majority (19; 63%) of the 30 autopsies revealed no significant fetal abnormalities except for intrauterine hypoxia and asphyxia. Among all 68 cases, SARS-CoV-2 was detected from a body specimen in 16 of 28 cases tested, most frequently from nasopharyngeal swabs. Four autopsied stillborns had SARS-CoV-2 identified in internal organs. CONCLUSIONS.­: The pathology abnormalities composing SARS-CoV-2 placentitis cause widespread and severe placental destruction resulting in placental malperfusion and insufficiency. In these cases, intrauterine and perinatal death likely results directly from placental insufficiency and fetal hypoxic-ischemic injury. There was no evidence that SARS-CoV-2 involvement of the fetus had a role in causing these deaths.

Decitabine potentiates efficacy of doxorubicin in a preclinical trastuzumab-resistant HER2-positive breast cancer models.

Acquired drug resistance and metastasis in breast cancer (BC) are coupled with epigenetic deregulation of gene expression. Epigenetic drugs, aiming to reverse these aberrant transcriptional patterns and sensitize cancer cells to other therapies, provide a new treatment strategy for drug-resistant tumors. Here we investigated the ability of DNA methyltransferase (DNMT) inhibitor decitabine (DAC) to increase the sensitivity of BC cells to anthracycline antibiotic doxorubicin (DOX). Three cell lines representing different molecular BC subtypes, JIMT-1, MDA-MB-231 and T-47D, were used to evaluate the synergy of sequential DAC + DOX treatment in vitro. The cytotoxicity, genotoxicity, apoptosis, and migration capacity were tested in 2D and 3D cultures. Moreover, genome-wide DNA methylation and transcriptomic analyses were employed to understand the differences underlying DAC responsiveness. The ability of DAC to sensitize trastuzumab-resistant HER2-positive JIMT-1 cells to DOX was examined in vivo in an orthotopic xenograft mouse model. DAC and DOX synergistic effect was identified in all tested cell lines, with JIMT-1 cells being most sensitive to DAC. Based on the whole-genome data, we assume that the aggressive behavior of JIMT-1 cells can be related to the enrichment of epithelial-to-mesenchymal transition and stemness-associated pathways in this cell line. The four-week DAC + DOX sequential administration significantly reduced the tumor growth, DNMT1 expression, and global DNA methylation in xenograft tissues. The efficacy of combination therapy was comparable to effect of pegylated liposomal DOX, used exclusively for the treatment of metastatic BC. This work demonstrates the potential of epigenetic drugs to modulate cancer cells' sensitivity to other forms of anticancer therapy.

Intrauterine Fetal Demise After Uncomplicated COVID-19: What Can We Learn from the Case?

BACKGROUND: SARS-CoV-2 infection in pregnant women can lead to placental damage and transplacental infection transfer, and intrauterine fetal demise is an unpredictable event. CASE STUDY: A 32-year-old patient in her 38th week of pregnancy reported loss of fetal movements. She overcame mild COVID-19 with positive PCR test 22 days before. A histology of the placenta showed deposition of intervillous fibrinoid, lympho-histiocytic infiltration, scant neutrophils, clumping of villi, and extant infarctions. Immunohistochemistry identified focal SARS-CoV-2 nucleocapsid and spike protein in the syncytiotrophoblast and isolated in situ hybridization of the virus' RNA. Low ACE2 and TMPRSS2 contrasted with strong basigin/CD147 and PDL-1 positivity in the trophoblast. An autopsy of the fetus showed no morphological abnormalities except for lung interstitial infiltrate, with prevalent CD8-positive T-lymphocytes and B-lymphocytes. Immunohistochemistry and in situ hybridization proved the presence of countless dispersed SARS-CoV-2-infected epithelial and endothelial cells in the lung tissue. The potential virus-receptor protein ACE2, TMPRSS2, and CD147 expression was too low to be detected. CONCLUSION: Over three weeks' persistence of trophoblast viral infection lead to extensive intervillous fibrinoid depositions and placental infarctions. High CD147 expression might serve as the dominant receptor for the virus, and PDL-1 could limit maternal immunity in placental tissue virus clearance. The presented case indicates that the SARS-CoV-2 infection-induced changes in the placenta lead to ischemia and consecutive demise of the fetus. The infection of the fetus was without significant impact on its death. This rare complication of pregnancy can appear independently to the severity of COVID-19's clinical course in the pregnant mother.

KIT Expression Is Regulated by DNA Methylation in Uveal Melanoma Tumors.

Uveal melanoma (UM) is an ocular tumor with a dismal prognosis. Despite the availability of precise molecular and cytogenetic techniques, clinicopathologic features with limited accuracy are widely used to predict metastatic potential. In 51 UM tissues, we assessed a correlation between the expression of nine proteins evaluated by immunohistochemistry (IHC) (Melan-A, S100, HMB45, Cyclin D1, Ki-67, p53, KIT, BCL2, and AIFM1) and the presence of UM-specific chromosomal rearrangements measured by multiplex ligation-dependent probe amplification (MLPA), to find IHC markers with increased prognostic information. Furthermore, mRNA expression and DNA methylation values were extracted from the whole-genome data, achieved by analyzing 22 fresh frozen UM tissues. KIT positivity was associated with monosomy 3, increasing the risk of poor prognosis more than 17-fold (95% CI 1.53-198.69, p = 0.021). A strong negative correlation was identified between mRNA expression and DNA methylation values for 12 of 20 analyzed positions, five located in regulatory regions of the KIT gene (r = -0.658, p = 0.001; r = -0.662, p = 0.001; r = -0.816; p < 0.001; r = -0.689, p = 0.001; r = -0.809, p < 0.001, respectively). DNA methylation ß values were also inversely associated with KIT protein expression (p = 0.001; p = 0.001; p = 0.015; p = 0.025; p = 0.002). Our findings, showing epigenetic deregulation of KIT expression, may contribute to understanding the past failure to therapeutically target KIT in UM.

Chemotherapy-triggered changes in stromal compartment drive tumor invasiveness and progression of breast cancer.

BACKGROUND: Chemotherapy remains a standard treatment option for breast cancer despite its toxic effects to normal tissues. However, the long-lasting effects of chemotherapy on non-malignant cells may influence tumor cell behavior and response to treatment. Here, we have analyzed the effects of doxorubicin (DOX) and paclitaxel (PAC), commonly used chemotherapeutic agents, on the survival and cellular functions of mesenchymal stromal cells (MSC), which comprise an important part of breast tumor microenvironment. METHODS: Chemotherapy-exposed MSC (DOX-MSC, PAC-MSC) were co-cultured with three breast cancer cell (BCC) lines differing in molecular characteristics to study chemotherapy-triggered changes in stromal compartment of the breast tissue and its relevance to tumor progression in vitro and in vivo. Conditioned media from co-cultured cells were used to determine the cytokine content. Mixture of BCC and exposed or unexposed MSC were subcutaneously injected into the immunodeficient SCID/Beige mice to analyze invasion into the surrounding tissue and possible metastases. The same mixtures of cells were applied on the chorioallantoic membrane to study angiogenic potential. RESULTS: Therapy-educated MSC differed in cytokine production compared to un-exposed MSC and influenced proliferation and secretory phenotype of tumor cells in co-culture. Histochemical tumor xenograft analysis revealed increased invasive potential of tumor cells co-injected with DOX-MSC or PAC-MSC and also the presence of nerve fiber infiltration in tumors. Chemotherapy-exposed MSC have also influenced angiogenic potential in the model of chorioallantoic membrane. CONCLUSIONS: Data presented in this study suggest that neoadjuvant chemotherapy could possibly alter otherwise healthy stroma in breast tissue into a hostile tumor-promoting and metastasis favoring niche. Understanding of the tumor microenvironment and its complex net of signals brings us closer to the ability to recognize the mechanisms that prevent failure of standard therapy and accomplish the curative purpose.

Hyperbilirubinemia Maintained by Chronic Supplementation of Unconjugated Bilirubin Improves the Clinical Course of Experimental Autoimmune Arthritis.

Rheumatoid arthritis (RA) is a chronic multisystem disease, therapy of which remains a challenge for basic research. The present work examined the effect of unconjugated bilirubin (UCB) administration in adjuvant-induced arthritis (AIA)-an experimental model, in which oxidative stress (OS), inflammation and inadequate immune response are often similar to RA. Male Lewis rats were randomized into groups: CO-control, AIA-untreated adjuvant-induced arthritis, AIA-BIL-adjuvant-induced arthritis administrated UCB, CO-BIL-control with administrated UCB. UCB was administered intraperitoneally 200 mg/kg of body weight daily from 14th day of the experiment, when clinical signs of the disease are fully manifested, to 28th day, the end of the experiment. AIA was induced by a single intradermal immunization at the base of the tail with suspension of Mycobacterium butyricum in incomplete Freund's adjuvant. Clinical, hematologic, biochemical and histologic examinations were performed. UCB administration to animals with AIA lead to a significant decrease in hind paws volume, plasma levels of C-reactive protein (CRP) and ceruloplasmin, drop of leukocytes, lymphocytes, erythrocytes, hemoglobin and an increase in platelet count. UCB administration caused significantly lowered oxidative damage to DNA in arthritic animals, whereas in healthy controls it induced considerable oxidative damage to DNA. UCB administration also induced atrophy of the spleen and thymus in AIA and CO animals comparing to untreated animals. Histological signs of joint damage assessed by neutrophils infiltration and deposition of fibrin were significantly reduced by UCB administration. The effects of exogenously administered UCB to the animals with adjuvant-induced arthritis might be identified as therapeutic, in contrast to the effects of UCB administration in healthy animals rather classified as toxic.

Prognostic Value of Apoptosis-Inducing Factor (AIF) in Germ Cell Tumors.

Apoptosis is a strictly regulated process essential for preservation of tissue homeostasis. This study aimed to evaluate expression of apoptosis inducing factor (AIF) in testicular germ cell tumors (GCTs) and to correlate expression patterns with clinicopathological variables. Formalin-fixed and paraffin-embedded specimens of non-neoplastic testicular tissue and GCTs obtained from 216 patients were included in the study. AIF expression was detected by immunohistochemistry, scored by the multiplicative quickscore method (QS). Normal testicular tissue exhibits higher cytoplasmic granular expression of AIF compared to GCTs (mean QS = 12.77 vs. 4.80, p < 0.0001). Among invasive GCTs, mean QS was the highest in embryonal carcinoma, yolk sac tumor and seminoma, lower in teratoma and the lowest in choriocarcinoma. No nuclear translocation of AIF was observed. Nonpulmonary visceral metastases were associated with lower AIF expression. Metastatic GCTs patients with high AIF expression had better overall survival compared to patients with low AIF expression (HR = 0.26, 95% CI 0.11-0.62, p = 0.048). We observed significantly lower AIF expression in GCTs compared to normal testicular tissue, which is an uncommon finding in malignant tumors. AIF downregulation might represent one of the mechanisms of inhibition of apoptosis and promotion of cell survival in GCTs.

Immunohistochemical evaluation of potential molecular targets of desmoid-type fibromatosis.

Desmoid-type fibromatosis is locally aggressive tumor rare in general population, although commonly present in patients with familial adenomatous polyposis, significantly contributing to the morbidity and mortality of patients. To optimize and individualize the management of patients it is necessary to better understand the biology of these tumors. Immunohistochemical analysis of ß-catenin, VEGF, hormone receptors ERß, ERα and PR, COX-2, APC protein, EGFR, c-kit (CD117), bcl-2 and HER2 expression, potential therapeutic targets, was carried out on 15 archival biopsy samples together with APC gene mutational screening. ß-catenin expression was found in all samples, with over 73% showing high range positivity, however with no prognostic significance. Non-specific cytoplasmic localization of ß-catenin was observed FAP-associated cases lacking CTNNB1 mutations. Hormone receptor status demonstrated expression of ERß in 93% of lesions, without detectable ERα or PR. Distinct COX-2 expression of variable intensity was present in all but one desmoid-type fibromatosis case. All lesions demonstrated intense VEGF positivity. Immunoreactivity for the APC protein was found only in 4 cases associated with FAP. No EGFR, HER2, bcl-2 or c-kit expression was detected in any sample. Expression of ß-catenin, VEGF, ERß, COX-2 in high number of cases suggests a potential as future therapeutic targets in desmoid-type fibromatosis.

Monosomy 3 Influences Epithelial-Mesenchymal Transition Gene Expression in Uveal Melanoma Patients; Consequences for Liquid Biopsy.

Despite outstanding advances in diagnosis and the treatment of primary uveal melanoma (UM), nearly 50% of UM patients develop metastases via hematogenous dissemination, driven by the epithelial-mesenchymal transition (EMT). Despite the failure in UM to date, a liquid biopsy may offer a feasible non-invasive approach for monitoring metastatic disease progression and addressing protracted dormancy. To detect circulating tumor cells (CTCs) in UM patients, we evaluated the mRNA expression of EMT-associated transcription factors in CD45-depleted blood fraction, using qRT-PCR. ddPCR was employed to assess UM-specific GNA11, GNAQ, PLCß4, and CYSLTR2 mutations in plasma DNA. Moreover, microarray analysis was performed on total RNA isolated from tumor tissues to estimate the prognostic value of EMT-associated gene expression. In total, 42 primary UM and 11 metastatic patients were enrolled. All CD45-depleted samples were negative for CTC when compared to the peripheral blood fraction of 60 healthy controls. Tumor-specific mutations were detected in the plasma of 21.4% patients, merely, in 9.4% of primary UM, while 54.5% in metastatic patients. Unsupervised hierarchical clustering of differentially expressed EMT genes showed significant differences between monosomy 3 and disomy 3 tumors. Newly identified genes can serve as non-invasive prognostic biomarkers that can support therapeutic decisions.