Folia Biologica最新文献

Transient receptor potential cation channel subfamily V member 1 (TRPV1) has been found over-expressed in low back pain (LBP) patients with neuropathic pain (NP), but the underlying mechanism is still unclear. In the present study, the up-regulation of the TRPV1 protein level in sinuvertebral nerve biopsies from patients with NP was verified by immunoblotting, but the TRPV1 mRNA level was not significantly changed. MiRNAs targeting TRPV1 mRNA were predicted by a bioinformatic tool, and the interactions between the miRNAs and TRPV1 were confirmed by dual luciferase assay. The correlation between NFKB1 signalling and TRPV1 expression was analysed and confirmed by using sNF96.2 cells after lipopolysaccharide stimulation. We found that five out of 18 miRNAs repressed TRPV1 expression, and the levels of miR-375 and miR-455 were negatively correlated with the protein level of TRPV1 in patients with NP. MiR-375 and miR-455 were identified to repress TRPV1 expression via targeting the 3'UTR of TRPV1 mRNA. NFKB1 signalling activation down-regulated the expression of miR-375 and miR-455, and thus up-regulated the TRPV1 protein level. In conclusion, we partially unveiled the mechanism of how TRPV1 is over-expressed in chronic LBP patients with NP and provided two potential candidate miRNAs for NP treatment.

We compared the efficiency of real-time PCR analysis of FII (c.*97G>A, G20210A) and FV Leiden (c.1601G>A) thrombophilic mutations in the samples obtained from venous blood treated with various anti coagulant agents (EDTA, heparin, and sodium fluoride with potassium oxalate), or from clotted venous blood; one hundred samples of wild-type subjects were tested. Genomic DNA extracts and whole blood specimens modified by 90 °C heating were analysed by real-time PCR analysis; cycle threshold values were subsequently evaluated. Real-time PCR analysis for the FII gene assay performed in DNA extracts from EDTA blood samples revealed a median Ct value of 19.3. Similar Ct values were apparent in the DNA extracts obtained from the heparinized blood and sodium fluoride with potassium oxalatetreated samples: 18.5 and 18.9, respectively. Significantly higher Ct values were found in extracts from clotted blood with medians of 20.6 (tubes with inert separation gel) and 20.5 (tubes without the gel, both P < 0.001). The data on the FV real-time PCR analysis were very comparable to the FII assay. In the modified whole blood, the samples treated with heparin salts showed significantly lower Ct values (P < 0.001) in both assays when compared with the samples with EDTA, sodium fluoride with potassium oxalate, and with the samples with clotted blood. Our results indicate that real-time PCR analyses of thrombophilic mutations were not negatively influenced by the presence of heparin salts in collection tubes. Blood samples with various anticoagulants might be exchangeable for each other when DNA analysis of thrombophilic mutations is required.

Nucleolar RNA optical density (concentration) measurements at the single cell level indicated that differentiation of lymphocytes is accompanied by a slightly decreased nucleolar RNA concentration in contrast to the cytoplasmic rim around the nucleus. On the other hand, the nucleolar size was markedly reduced and the cytoplasmic rim surrounding the nucleus was reduced only weakly. Concerning the calculated rough estimate of the RNA content, the differentiation induced its larger decrease in the nucleoli than in the cytoplasmic rim. These observations indicated that the nucleolar RNA concentration and RNA content together with the nucleolar morphology are more sensitive markers of the differentiation process than the RNA concentration and content in the cytoplasm. Thus, the nucleolar RNA transfer to the cytoplasm in advanced differentiation steps might still be going on regardless of the decreasing or inhibited nucleolar biosynthetic activity. In addition, the presence of ring-shaped nucleoli and micronucleoli characteristic of mature and terminal lymphocytes in some lymphocytic less differentiated steps, i.e., lymphoblasts and prolymphocytes, might indicate the premature differentiation state of such cells.

Clear cell renal cell carcinoma (ccRCC) is very common and accounts for most kidney cancer deaths. While many studies are being conducted in finding the prognostic signatures of ccRCC, we believe that ferroptosis, which involves programmed cell death dependent on iron accumulation, has therapeutic potential in ccRCC. Recent research has shown that long noncoding RNAs (lncRNAs) are involved in ferroptosis-related tumour processes and are closely related to survival in patients with ccRCC. Hence, in this study we aim to further explore the role of ferroptosis-related lncRNAs (FRLs) in ccRCC, hoping to establish a signature to predict the survival outcome of ccRCC. We analysed transcriptome data from The Cancer Genome Atlas database (TCGA) and ferroptosis-related genes (FRGs) from FerrDb to identify FRLs using Pearson's correlation. Lasso Cox regression analysis and multivariate Cox proportional hazards models screened seventeen optimal FRLs for developing prognostic signatures. Kaplan-Meier survival curves and ROC curves were then plotted for validating the sensitivity, specificity, and accuracy of the identified signatures. Gene Set Enrichment Analysis and CIBERSORT algorithm were deployed to explore the role of these FRLs in the tumour microenvironment. It was concluded that these models demonstrate excellent performance in predicting prognosis among patients with ccRCC, also indicating association with the clinicopathologic parameters such as tumour grade, tumour stage and tumour immune infiltration. In conclusion, our findings provide novel insights into ferroptosis-related lncRNAs in ccRCC, which are important targets for investigating the tumorigenesis of ccRCC.

This is the first histological and molecular analysis of two chondrosarcomas with target-like chondrocytes that were compared with a group of conventional chondrosarcomas and enchondromas. The unique histological feature of target-like chondrocytes is the presence of unusual hypertrophic eosinophilic APAS-positive perichondrocytic rings (baskets). In the sections stained with Safranin O/Fast green, the outer part of the ring was blue and the material in the lacunar space stained orange, similarly to intercellular regions. Immunohistochemical examination showed strong positivity for vimentin, factor XIIIa, cyclin D1, osteonectin, B-cell lymphoma 2 apoptosis regulator (Bcl-2), p53 and p16. The S-100 protein was positive in 25 % of neoplastic cells. Antibodies against GFAP, D2-40 (podoplanin), CD99, CKAE1.3 and CD10 exhibited weak focal positivity. Pericellular rings/baskets contained type VI collagen in their peripheral part, in contrast to the type II collagen in intercellular interterritorial spaces. Ultrastructural examination revealed that pericellular rings contained an intralacunar component composed of microfibrils with abundant admixture of aggregates of dense amorphous non-fibrillar material. The outer extralacunar zone was made up of a layer of condensed thin collagen fibrils with admixture of non-fibrillar dense material. NGS sequencing identified a fusion transcript involving fibronectin 1 (FN1) and fibroblast growth factor receptor 2 (FGFR2) at the RNA level. At the DNA level, no significant variant was revealed except for the presumably germline variant in the SPTA1 gene.

Diarrhoea is a common clinical condition; its pathogenesis is strongly associated with gut microbiota dysbiosis. Limonitum is a well-known traditional Chinese medicine that exerts appreciable benefits regarding the amelioration of diarrhoea. However, the mechanism through which Limonitum ameliorates diarrhoea remains unclear. Here, the efficacy and underlying mechanism of Limonitum decoction (LD) regarding diarrhoea were explored from the aspect of gut microbiota. Castor oil (CO) was used to induce diarrhoea in mice, which were then used to evaluate the effects of LD regarding the timing of the first defecation, diarrhoea stool rate, degree of diarrhoea, diarrhoea score, intestinal propulsive rate, and weight of intestinal contents. The concentrations of short-chain fatty acids (SCFAs), including acetic, propionic, isobutyric, butyric and valeric acids, were analysed by gas chromatography-mass spectrometry (GC-MS). The 16S rRNA high-throughput sequencing technology was applied to evaluate changes in the gut microbiota under exposure to LD. LD was found to effectively ameliorate the symptoms of diarrhoea, and the diversity and relative abundance of gut microbiota were restored to normal levels following LD treatment. Additionally, LD significantly restored the observed reductions in SCFAs. These results provide strong evidence that LD can sufficiently ameliorate diarrhoea in mice by regulating their gut microbiota. The findings presented here highlight that Limonitum may constitute a prospective remedy for diarrhoea.

As the number of cancer patients globally increases, a need for reliable biomarkers including circulating tumour DNA from liquid biopsy for diagnosis, prognosis and monitoring of the disease is rising. Currently, mainly tissue samples from biopsy are used, but there are certain limitations: firstly, it is an invasive technique, and secondly, in some cases it is almost impossible to obtain an acceptable tissue sample. This could be changed by using circulating cell-free DNA from liquid biopsy, which also gives the possibility of repeated examination. Here, we focus on the options of isolating circulating cell-free DNA from plasma samples using two isolation techniques: precision manual QIAamp Circulating Nucleic Acid Kit and automatic MagNA Pure Compact (MPC) using Nucleic Acid Isolation Kit I. Manual extraction gave significantly better yields of circulating tumour DNA (P < 0.05). This DNA also had less contaminants (organic compounds or proteins). DNA obtained by both tested methods of isolation is suitable for subsequent molecular genetic methods.

Recently, more and more efforts are directed towards developing new imaging and drug-delivery options based on various nanoparticles, exploiting their unique properties. Here, ultra-small gold nanoparticles functionalized with widely used polyethylene glycol and its amine-terminated form were tested in respect of their potential interactions with human immune cells (cell line and primary cells). The results showed that differently terminated ultrasmall gold nanoparticles represent an interesting theranostic platform as they are harmless to immune cells (not inducing cytotoxicity and severe immune response) and on the other hand, they can serve as imaging and/or drug delivery agents using e.g. monocytes/ macrophages as "Trojan horses" to deliver these nanoparticles across the blood-brain barrier and diagnose or treat pathologies of the central nervous system.

Proliferation and migration of retinal endothelial cells (RECs) contribute to the development of diabetic retinopathy. PLAG1 (pleomorphic adenoma gene 1) functions as a zinc-finger transcription factor to participate in the development of lipoblastomas or pleomorphic adenomas of the salivary glands through regulation of cell proliferation and migration. The role of PLAG1 in diabetic retinopathy was investigated in this study. Firstly, RECs were induced under high glucose conditions, which caused reduction in viability and induction of apoptosis in the RECs. Indeed, PLAG1 was elevated in high glucosetreated RECs. Functional assays showed that silence of PLAG1 increased viability and suppressed apoptosis in high glucose-induced RECs, accompanied with up-regulation of Bcl-2 and down-regulation of Bax and cleaved caspase-3. Moreover, migration of RECs was promoted by high glucose conditions, while repressed by knockdown of PLAG1. High glucose also triggered angiogenesis of RECs through up-regulation of vascular endothelial growth factor (VEGF). However, interference of PLAG1 reduced VEGF expression to retard the angiogenesis. Silence of PLAG1 also attenuated high glucose-induced up-regulation of Wnt3a, β-catenin and c-Myc in RECs. Moreover, silence of PLAG1 ameliorated histopathological changes in the retina of STZ-induced diabetic rats through down-regulation of β-catenin. In conclusion, knockdown of PLAG1 suppressed high glucose-induced angiogenesis and migration of RECs, and attenuated diabetic retinopathy by inactivation of Wnt/ β-catenin signalling.

Autologous serum eye drops (ASEDs) are used as a treatment for severe dry eye disease. The concentration and stability of various growth factors in ASEDs is determinative for their efficiency. We therefore assessed the concentrations of transforming growth factor beta 1 (TGF-β1), epidermal growth factor (EGF) and insulin-like growth factor 1 (IGF-1) in ASEDs following storage at 4-8, -20, -80 and -156 °C. Twenty % and 100% sera from eight healthy volunteers were analysed by the sandwich enzyme immunoassay at different time intervals up to seven months. The mean levels of TGF-β1 and EGF in undiluted and 20% serum did not differ significantly from the baseline levels in fresh serum for any storage conditions after 7 days at 4-8 °C, as well as after 4- and 7-month preservation at sub-zero temperatures. In 20% serum, no IGF-1 concentration decrease was found following 7 days of preservation at 4-8 °C. However, a decrease to 78 % and 81 % (P < 0.01) of baseline values was found in 20% serum after 4-month storage at -20 °C and 7-month storage at -156 °C, respectively. A more pronounced decrease in IGF-1 was observed in undiluted serum. All assessed growth factors present in 20% frozen serum remained stable for up to 7 months. The highest stability was achieved at -80 °C. At -20 and -156 °C, some decrease in IGF-1 occurred. Our results indicate that 20% ASEDs can be stored frozen up to 7 months under proper conditions.