miRNA expression in triple-negative breast cancers (TNBC) has mainly been studied from a methodological viewpoint. However, it has not been considered that miRNA expression profile may be associated with a specific morphological entity inside every tumor. The verification of this hypothesis on a set of 25 TNBCs was the subject of our previous work, where we confirmed specific expression of the studied miRNAs in 82 samples of different morphologies including inflammatory infiltrate, spindle cell, clear cell, and metastases after RNA extraction and purification as well as microchip and biostatistical analysis. In the current work, we demonstrate a low suitability of in situ hybridization method for miRNA detection compared to RT-qPCR, and in detail discuss the biological role of 8 miRNAs with the most significant changes of expression.
- MeSH
- lidé MeSH
- mikro RNA * genetika metabolismus MeSH
- triple-negativní karcinom prsu * genetika patologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
AIMS: The aim of this study was to compare the expression profile of selected DNA methyltransferases and global DNA methylation status in patients with different phases of multiple myeloma (MM) . For the analysis, different cellular populations including unsorted myeloma cells and a set of plasma cells purified by relevant antibodies were used. Consequently, laboratory data were compared to patients' clinical data. PATIENTS AND METHODS: For the analysis, unsorted bone marrow cell population of 44 MM patients (30 newly diagnosed, 9 relapsed and 5 patients in remission) and a set of 8 patients' samples of sorted plasma cells were used. We used commercially available RNA isolated from BM of 3 healthy individuals as control samples. Expression analysis of three DNA methyltransferases - DNMT1, DNMT3A, and DNMT3B was performed by quantitative RT-PCR and the patient global DNA methylation profiles were detected by colorimetric assay. RESULTS: Unchanged DNMT1 expression was detected in the selected cohort of patients. Normalized DNMT3A gene expression was globally higher in comparison with controls in unsorted and sorted cell populations. Low (0.08-1.81%) global DNA methylation status in unsorted samples of multiple myeloma patients did not correlate either with expression profiles of monitored DNA methyltransferases or with the stages of MM based on Durie-Salmon and International Staging System. CONCLUSION: This is the first comparative study between DNA methyltransferases expression profiles and global DNA methylation status in different phases of multiple myeloma patients. No significant correlation between the level of global methylation and the clinical stage of the unsorted cell population of patients with multiple myeloma was registered. Overexpression of the DNMT3A gene occurred in both sorted and unsorted cell populations of patients with multiple myeloma. This fact highlights the DNMT3A as a potential marker of multiple myeloma tumor progression. Moreover, we demonstrated comparable results in the expression of DNA methyltransferases in both sorted and unsorted cell populations. This is a promising result from the methodical point of view because when compared to samples of unsorted multiple myeloma cells, samples of sorted cells bring reduction of the number of possible analyses performed.
- MeSH
- DNA methyltransferasa 3A MeSH
- DNA MeSH
- lidé MeSH
- metylace DNA * MeSH
- mnohočetný myelom * genetika MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Východiska: Současné in vitro modelové systémy plně neodrážejí biologickou a klinickou diverzitu karcinomu prostaty (prostate cancer – PCa). Organoidy jsou 3D in vitro buněčné kultury, které mohou lépe rekapitulovat heterogenitu onemocnění a zachovat vlastnosti původního nádoru. Krátkodobá ex vivo kultivace PCa tkání může také usnadnit testování léčiv v personalizované medicíně. Materiál a metody: Pro organoidní kultivaci jsme zpracovali jak nádorovou, tak normální tkáň od 50 pacientů, kteří podstoupili radikální prostatektomii nebo transuretrální resekci prostaty. Kromě toho jsme využili techniku ex vivo tkáňové kultivace a provedli krátkodobý experiment s použitím gemcitabinu a inhibitoru Chk1 MU380 ve vzorcích od 10 pacientů. Výsledky: Celkem jsme byli schopni kultivovat organoidy z 58 % nádorových (29/50) a 69 % normálních tkání (20/29). Imunohistochemické barvení dvou reprezentativních případů odhalilo buněčnou pozitivitu na pan-cytokeratin potvrzující přítomnost epiteliálních buněk. Nadměrná exprese proteinů AMACR a ERG v nádorech však nebyla zachována v organoidech. Dalším omezením bylo udržení organoidů pouze do první pasáže, obvykle po dobu 3 týdnů. Dále byly provedeny krátkodobé testy v ex vivo kultuře nádorových tkání od deseti pacientů. Tkáňové vzorky z prostatektomií většinou vykazovaly nízkou míru proliferace a Ki-67 pozitivity. Další nevýhodou tohoto přístupu byla nekonzistentní morfologie mezi jednotlivými tkáňovými fragmenty. Pouze jeden případ vykazoval vysokou míru proliferace pro testování léčiv a nádorová tkáň byla přítomna ve všech testovaných vzorcích. V naší práci také poskytujeme přehled nedávných studií a podrobné srovnání kultivačních podmínek. Závěr: Podařilo se nám ustavit kultury organoidů i fragmentů tkání z primárních nádorů prostaty. Exprese nádorových markerů však nebyla zachována v získaných organoidech. Nekonzistentní morfologie a nízká proliferace ztěžovaly interpretaci výsledků testování léčiv u většiny případů. Přesto mohou být tyto přístupy slibné při použití tkání z metastatického kastračně rezistentního karcinomu prostaty.
Background: Current in vitro model systems do not fully reflect the biological and clinical diversity of prostate cancer (PCa). Organoids are 3D in vitro cell cultures that may better recapitulate disease heterogeneity and retain parental tumor characteristics. Short-term ex vivo culture of PCa tissues may also facilitate drug testing in personalized medicine. Materials and methods: For organoid culture, we have processed both cancer and normal tissues from 50 patients who underwent radical prostatectomy or transurethral resection of the prostate. In addition, we exploited the ex vivo tissue culture technique and performed short-term chemotherapy assay using gemcitabine and Chk1 inhibitor MU380 in 10 patient samples. Results: In total, we were able to cultivate organoids from 58% of tumors (29/50) and 69% of normal tissue (20/29). Immunohistochemical staining of two representative cases revealed cell positivity for pan-cytokeratin confirming the presence of epithelial cells. However, the overexpression of AMACR and ERG proteins in tumors was not recapitulated in organoids. Another limitation was the propagation of organoids only up to 3 weeks till the first passage. Next, a short-term drug test was performed for ten patients using ex vivo tissue culture. Samples from prostatectomies mostly presented a low proliferation rate as assessed by Ki-67 staining. Another drawback of this approach was inconsistent tissue morphology among particular tissue fragments. Only one case showed a high proliferation rate for drug testing and tumor tissue was present in all tested samples. In our work, we also provide an overview of recent studies and a detailed comparison of culture conditions. Conclusion: We have established cultures of both organoids and tissue fragments from PCa patient samples. However, the expression of tumor markers was not recapitulated in organoids. Inconsistent morphology among tissue fragments and low proliferation hampered the interpretation of the drug testing in most cases. Still, these approaches may be promising using tissues from metastatic castration-resistant prostate cancer.
- Klíčová slova
- ex vivo,
- MeSH
- individualizovaná medicína metody MeSH
- klinická studie jako téma MeSH
- kultivační techniky MeSH
- lidé MeSH
- nádorové buňky kultivované MeSH
- nádory prostaty * patologie MeSH
- organoidy metabolismus patologie MeSH
- transuretrální resekce prostaty MeSH
- Check Tag
- lidé MeSH
- mužské pohlaví MeSH
- Publikační typ
- práce podpořená grantem MeSH
BACKGROUND: Meningioma is the most common primary central nervous system neoplasm, accounting for about a third of all brain tumors. Because their growth rates and prognosis cannot be accurately estimated, biomarkers that enable prediction of their biological behavior would be clinically beneficial. OBJECTIVE: To identify coding and noncoding RNAs crucial in meningioma prognostication and pathogenesis. METHODS: Total RNA was purified from formalin-fixed and paraffin-embedded tumor samples of 64 patients with meningioma with distinct clinical characteristics (16 recurrent, 30 nonrecurrent with follow-up of >5 years, and 18 with follow-up of <5 years without recurrence). Transcriptomic sequencing was performed using the HiSeq 2500 platform (Illumina), and biological and functional differences between meningiomas of different types were evaluated by analyzing differentially expression of messenger RNA (mRNA) and long noncoding RNA (IncRNA). The prognostic value of 11 differentially expressed RNAs was then validated in an independent cohort of 90 patients using reverse transcription quantitative (real-time) polymerase chain reaction. RESULTS: In total, 69 mRNAs and 108 lncRNAs exhibited significant differential expression between recurrent and nonrecurrent meningiomas. Differential expression was also observed with respect to sex (12 mRNAs and 59 lncRNAs), World Health Organization grade (58 mRNAs and 98 lncRNAs), and tumor histogenesis (79 mRNAs and 76 lncRNAs). Lnc-GOLGA6A-1, ISLR2, and AMH showed high prognostic power for predicting meningioma recurrence, while lnc-GOLGA6A-1 was the most significant factor for recurrence risk estimation (1/hazard ratio = 1.31; P = .002). CONCLUSION: Transcriptomic sequencing revealed specific gene expression signatures of various clinical subtypes of meningioma. Expression of the lnc-GOLGA61-1 transcript was found to be the most reliable predictor of meningioma recurrence.
- MeSH
- lidé MeSH
- lokální recidiva nádoru * diagnóza genetika MeSH
- meningeální nádory * diagnóza genetika MeSH
- meningeom * diagnóza genetika MeSH
- prognóza MeSH
- RNA dlouhá nekódující * genetika metabolismus MeSH
- stanovení celkové genové exprese MeSH
- transkriptom MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Mucosal surfaces are colonized by highly diverse commensal microbiota. Coating with secretory IgA (SIgA) promotes the survival of commensal bacteria while it inhibits the invasion by pathogens. Bacterial coating could be mediated by antigen-specific SIgA recognition, polyreactivity, and/or by the SIgA-associated glycans. In contrast to many in vitro studies, only a few reported the effect of SIgA glycans in vivo. Here, we used a germ-free antibody-free newborn piglets model to compare the protective effect of SIgA, SIgA with enzymatically removed N-glycans, Fab, and Fc containing the secretory component (Fc-SC) during oral necrotoxigenic E. coli O55 challenge. SIgA, Fab, and Fc-SC were protective, whereas removal of N-glycans from SIgA reduced SIgA-mediated protection as demonstrated by piglets' intestinal histology, clinical status, and survival. In vitro analyses indicated that deglycosylation of SIgA did not reduce agglutination of E. coli O55. These findings highlight the role of SIgA-associated N-glycans in protection. Further structural studies of SIgA-associated glycans would lead to the identification of those involved in the species-specific inhibition of attachment to corresponding epithelial cells.
- MeSH
- aglutinace MeSH
- Escherichia coli fyziologie MeSH
- glykosylace MeSH
- gnotobiologické modely MeSH
- imunoglobulin A sekreční metabolismus MeSH
- imunoglobuliny - Fab fragmenty metabolismus MeSH
- infekce vyvolané Escherichia coli imunologie MeSH
- jednořetězcové protilátky metabolismus MeSH
- novorozená zvířata MeSH
- odolnost vůči nemocem MeSH
- polysacharidy metabolismus MeSH
- prasata MeSH
- těhotenství MeSH
- zvířata MeSH
- Check Tag
- těhotenství MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND/AIM: Multiple myeloma is a highly heterogeneous disease of clonal plasma cells. Histone deacetylase (HDAC) inhibitors are promising anticancer drugs but their precise mechanisms of actions are not well understood. MATERIALS AND METHODS: Cell-cycle regulation and pro-apoptotic effects of two histone deacetylase inhibitors, suberohydroxamic acid (SAHA) and suberoylanilide hydroxamic acid (SBHA), were analyzed in multiple myeloma cell lines RPMI8226 and U266 with differing TP53 status using gene-expression analysis. RESULTS: Enhanced expression of cyclin-dependent kinase inhibitor 1A (CDKN1A/p21WAF/CIP1) detected in the TP53-deleted U266 cell line after SAHA treatment indicates the P53-independent mode of transcriptional activation of CDKN1A gene. In contrast, CDKN1A gene expression was significantly increased by both SBHA and SAHA treatment of TP53-mutated RPMI8226 cells. CONCLUSION: SAHA appears to be a potentially effective pro-apoptotic and anticancer drug with universal application in the treatment of heterogeneous populations of multiple myeloma cells.
- MeSH
- apoptóza účinky léků MeSH
- inhibitor p21 cyklin-dependentní kinasy antagonisté a inhibitory genetika MeSH
- inhibitory histondeacetylas farmakologie MeSH
- kontrolní body buněčného cyklu účinky léků MeSH
- kyseliny hydroxamové farmakologie MeSH
- lidé MeSH
- mnohočetný myelom farmakoterapie genetika metabolismus patologie MeSH
- nádorové buněčné linie MeSH
- nádorový supresorový protein p53 genetika metabolismus MeSH
- protinádorové látky farmakologie MeSH
- regulace genové exprese u nádorů účinky léků MeSH
- viabilita buněk účinky léků MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
OBJECTIVES: DNA repair proteins have emerged as potential predictors for immunotherapy response alongside PD-L1 expression, tumor-infiltrating lymphocytes (TILs) and tumor mutational burden. We analyzed expression of PD-L1, TILs count and expression of the homologous recombination (HR) protein RAD51, as potential prognostic factors in patients with resected non-small-cell lung carcinoma (NSCLC). MATERIALS AND METHODS: Discovery set included 96 NSCLC patients from the University Hospital Olomouc (Czech Republic) and a replication set included 1109 NSCLC patients from University Hospital Zurich (Switzerland). Tissue microarrays (TMAs) were stained using the automated staining platform Ventana Benchmark Ultra with antibodies against RAD51,CD3, CD8, CD68 and PD-L1. RESULTS: Loss of nuclear RAD51 protein was associated with high TILs (r=-0.25, p = 0.01) and PD-L1 status (10.6 vs. 2.4 %, p = 0.012) in patients receiving neoadjuvant chemo-/radiotherapy (CT/RT). In silico analysis from the TCGA data set showed a negative relationship between RAD51 mRNA expression and CD45 (r = ‒0.422, p < 0.0001), CD68 (r = ‒0.326, p < 0.001), CD3 (r = ‒0.266, p < 0.001) and CD8 (r = ‒0.102, p < 0.001). RAD51 low/PD-L1 high patients were clustered as separate entity in the replication set and in TCGA dataset. High TILs status was significantly associated with improved OS in the replication set (unadjusted HR = 0.57, 95 % CI 0.42-0.76, p < 0.001). Similar results have been seen for CD3, CD8 and CD68. CONCLUSIONS: In conclusion, RAD51 nuclear loss is weakly associated with increased TILs and high PD-L1 at the time of surgery in curatively resected NSCLC and after prior exposure to neoadjuvant chemo- or radiotherapy. Both high TILs and RAD51 nuclear loss were confirmed as independent prognostic factors in curatively resected NSCLC.
- MeSH
- antigeny CD274 genetika MeSH
- lidé MeSH
- nádory plic * genetika terapie MeSH
- nemalobuněčný karcinom plic * genetika terapie MeSH
- oprava DNA MeSH
- prognóza MeSH
- rekombinasa Rad51 genetika MeSH
- tumor infiltrující lymfocyty MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Švýcarsko MeSH
BACKGROUND: Regional variability in dural sinus (DS) wall thickness in posterior cranial fossa (PCF) have not been studied in detail yet. OBJECTIVE: To clarify the possible regional variability in DS wall thickness and determine the occurrence and localization of the chordae Willisii (CW) in PCF. METHODS: Fifty-nine human cadaveric DSs of PCF were investigated. A measurement of the DS walls/dura mater/CW thickness of parafin-embedded/hematoxylin-eosin stained axial sections was performed by using Cell Sens Science Imaging Software (Olympus Corporation, Tokyo, Japan). RESULTS: The osseus wall (OW) was the thickest one in the confluens sinuum (CS) and the thinnest one in the jugular bulb (JB) and sigmoid sinus (P < .05). The biggest differences between individual walls were observed in the JB where the superior wall was almost twice as thick as the OW. At the transverse-sigmoid junction, the thickness of the walls was comparable. In the CS and transverse sinuses, the OW was even thicker than the surrounding dura mater. The occurrence and thickness of the CW increased from the JB towards CS and prevailed on the right side. An overall number of the CW in PCF was comparable to that observed in the superior sagittal sinus. CONCLUSION: The present study displayed for the first time the regional variability in the DS walls thickness and occurrence of the CW in PCF. Application of these findings may afford greater freedom in exposure of the DSs or neoplasms adhering to the DSs.
- MeSH
- dospělí MeSH
- dura mater * anatomie a histologie MeSH
- individualita MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mrtvola MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- sinus durae matris * anatomie a histologie MeSH
- zadní jáma lební * anatomie a histologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- abstrakt z konference MeSH
BACKGROUND: Regional variability in dural sinus (DS) wall thickness in posterior cranial fossa (PCF) have not been studied in detail yet. OBJECTIVE: To clarify the possible regional variability in DS wall thickness and determine the occurrence and localization of the chordae Willisii (CW) in PCF. METHODS: Fifty-nine human cadaveric DSs of PCF were investigated. A measurement of the DS walls/dura mater/CW thickness of parafin-embedded/hematoxylin-eosin stained axial sections was performed by using Cell Sens Science Imaging Software (Olympus Corporation, Tokyo, Japan). RESULTS: The osseus wall (OW) was the thickest one in the confluens sinuum (CS) and the thinnest one in the jugular bulb (JB) and sigmoid sinus (P < .05). The biggest differences between individual walls were observed in the JB where the superior wall was almost twice as thick as the OW. At the transverse-sigmoid junction, the thickness of the walls was comparable. In the CS and transverse sinuses, the OW was even thicker than the surrounding dura mater. The occurrence and thickness of the CW increased from the JB towards CS and prevailed on the right side. An overall number of the CW in PCF was comparable to that observed in the superior sagittal sinus. CONCLUSION: The present study displayed for the first time the regional variability in the DS walls thickness and occurrence of the CW in PCF. Application of these findings may afford greater freedom in exposure of the DSs or neoplasms adhering to the DSs.
- MeSH
- dospělí MeSH
- dura mater anatomie a histologie MeSH
- individualita MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mrtvola MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- sinus durae matris anatomie a histologie MeSH
- zadní jáma lební anatomie a histologie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
