Macrophage-Stimulating 1 Polymorphism rs3197999 in Pediatric Patients with Inflammatory Bowel Disease.

Background and Objectives: Inflammatory bowel disease (IBD), which includes Crohn's disease (CD) and ulcerative colitis (UC), often necessitates long-term treatment and hospitalizations and also may require surgery. The macrophage-stimulating 1 (MST1) rs3197999 polymorphism is strongly associated with the risk of IBD but its exact clinical correlates remain under investigation. We aimed to characterize the relationships between the MST1 rs3197999 genotype and the clinical characteristics in children and adolescents with IBD within a multi-center cross-sectional study. Materials and Methods: Clinical data included serum C-reactive protein (CRP), albumin, activity indices (PUCAI, PCDAI), anthropometric data, pharmacotherapy details, surgery, and disease severity. Genotyping for rs3197999 was carried out using TaqMan hydrolysis probes. Results: The study included 367 pediatric patients, 197 with Crohn's disease (CD) (40.6% female; a median age of 15.2 years [interquartile range 13.2-17.0]) and 170 with ulcerative colitis (UC) (45.8% female; a median age of 15.1 years [11.6-16.8]). No significant relationships were found between MST1 genotypes and age upon first biologic use, time from diagnosis to biological therapy introduction, PUCAI, PCDAI, or hospitalizations for IBD flares. However, in IBD, the height Z-score at the worst flare was negatively associated with the CC genotype (p = 0.016; CC: -0.4 [-1.2-0.4], CT: -0.1 [-0.7-0.8], TT: 0.0 [-1.2-0.7)]). The TT genotype was associated with higher C-reactive protein upon diagnosis (p = 0.023; CC: 4.3 mg/dL [0.7-21.8], CT 5.3 mg/dL [1.3-17.9], TT 12.2 mg/dL [3.0-32.9]). Conclusions: This study identified links between MST1 rs3197999 and the clinical characteristics of pediatric IBD: height Z-score and CRP. Further studies of the associations between genetics and the course of IBD are still warranted, with a focus on more extensive phenotyping.

Interleukin-6 as a Predictive Factor of Pathological Response to FLOT Regimen Systemic Treatment in Locally Advanced Gastroesophageal Junction or Gastric Cancer Patients.

BACKGROUND: Perioperative treatment is a gold standard in locally advanced gastric cancer or GEJ cancer in the Western population. Unfortunately, the response rate after neoadjuvant chemotherapy (NAC) remains limited. Moreover, there are currently no biomarkers enabling an individual prediction of therapeutic efficacy. The aim of this study was the identification of serum biomarkers of early response to NAC. METHODS: We conducted this prospective study in the MSCNRIO in Warsaw, Poland. A total of 71 patients and 15 healthy volunteers gave informed consent. Complete blood count, carcinoembryonic antigen (CEA), carcinoma antigen 125 (CA125), carcinoma antigen 19.9 (CA19.9), and fibrinogen (F) were measured at baseline and before every cycle. Circulating tumour cells (CTCs) and interleukin-1ß (IL-1ß), interleukin-6 (IL-6), interleukin-8 (IL-8), and interleukin-10 (IL-10) were measured in a pilot group of 40 patients at baseline and before cycle two (C2) and cycle three (C3). RESULTS: Of all the measured parameters, only the IL-6 serum level was statistically significant. The IL-6 level before C2 of chemotherapy was significantly decreased in the complete pathological response (pCR) vs. the non-pCR group (3.71 pg/mL vs. 7.63 pg/mL, p = 0.004). In all patients with an IL-6 level below 5.0 pg/mL in C2, tumour regression TRG1a/1b according to the Becker classification and ypN0 were detected in postoperative histopathological specimens. The IL-6 level before C1 of chemotherapy was significantly elevated in ypN+ vs. ypN0 (7.69 pg/mL vs. 2.89 pg/mL, p = 0.022). CONCLUSIONS: The trial showed that an elevated level of IL-6 prior to treatment and C2 might be a predictor of pathological response to NAC.

Development of Stable Infectious cDNA Clones of Tomato Black Ring Virus Tagged with Green Fluorescent Protein.

Tomato black ring virus (TBRV) is a member of the Nepovirus genus in the Secoviridae family, which infects a wide range of important crop species worldwide. In this work, we constructed four cDNA infectious clones of the TBRV tagged with the green fluorescent protein (TBRV-GFP), which varied in (i) the length of the sequences flanking the GFP insert, (ii) the position of the GFP insert within the RNA2 polyprotein, and (iii) the addition of a self-cutting 2A protein. The presence of the GFP coding sequence in infected plants was verified by RT-PCR, while the infectivity and stability of the constructs were verified by mechanical inoculation of the host plants. The systemic spread of TBRV-GFP within plants was observed under UV light at a macroscopic level, monitoring GFP-derived fluorescence in leaves, and at a microscopic level using confocal microscopy. The obtained clones are a valuable tool for future studies of TBRV-host interactions, virus biology, and the long-term monitoring of its distribution in infected plants.

Construction and biological characterization of a cDNA infectious clone of wheat umbra-like virus in wheat and Nicotiana benthamiana.

Umbravirus-like associated RNAs (ulaRNAs) are a new group of subviral RNAs associated with plants. Little is known about the biology of ulaRNAs. We recently reported wheat umbra-like virus (WULV) from Kansas fields. In this work, we generated a full-length cDNA clone of WULV which systemically infected N. benthamiana. While agroinfiltrated leaves demonstrated severe necrosis, upper leaves were symptomless. We also showed that WULV is capable of infecting wheat in the absence of a helper virus. Furthermore, and through sap inoculation, we demonstrated that WULV is transmissible in the form of free RNA. This is the first report of a mechanically transmissible ulaRNA. Together, these findings contribute to advancing our knowledge of the biology of WULV. Moreover, the construction of the WULV infectious clone provides a valuable research tool for further investigations including the role of WULV in symptom development in interaction with other wheat viruses.

C-Reactive Protein/Albumin Ratio at Diagnosis of Pediatric Inflammatory Bowel Disease: A Retrospective Multi-Center Study.

BACKGROUND This study aimed to evaluate the C-reactive protein-to-albumin (CRP/albumin) ratio at diagnosis of pediatric inflammatory bowel disease (IBD). MATERIAL AND METHODS Serum CRP/albumin ratio was calculated for patients with Crohn's disease (CD; n=186) and ulcerative colitis (UC; n=159) aged 3-18 years. RESULTS Patients with CD differed in CRP/albumin ratio at diagnosis in groups with quiescent, mild, moderate, and severe disease (P=0.011). CRP/albumin ratio at diagnosis was significant in differentiating patients with severe CD from quiescent disease at diagnosis (area under the curve (AUC)=0.94, odds ratio (OR)=63.4, 95% confidence interval (CI) 7.1-569.1, P<0.0001). CRP/albumin ratio at diagnosis could moderately differentiate penetrating from non-penetrating disease behavior in CD at diagnosis (AUC=0.73, OR=6.3, 95% CI 2.0-19.3, P<0.001). Furthermore, CRP/albumin ratio at diagnosis weakly differentiated IBD patients in need of biological treatment in a step-up procedure (AUC=0.58, OR=2.1, 95% CI 1.3-3.4, P=0.022) and in need of surgery (AUC=0.63, OR=3.1, 95% CI 1.4-7.2, P=0.006). For the IBD, CRP/albumin ratio at diagnosis was weakly correlated with age at first immunosuppressive treatment (rho=0.20, P=0.018), time from diagnosis to first biological treatment (rho=-0.37, P<0.001), days spent in hospital (rho=0.26, P=0.007), number of severe relapses (rho=0.31, P=0.001), and Pediatric Crohn's Disease Activity Index (rho=0.38, P=0.002). CONCLUSIONS The present findings add to previous studies carried out in adult patients and show that the CRP/albumin ratio at diagnosis was not significantly associated with the course of either CD or UC in children. However, CRP/albumin ratio could differentiate patients with severe CD from those with quiescent disease.

Genetic Diversity of Tomato Black Ring Virus Satellite RNAs and Their Impact on Virus Replication.

Viral satellite RNAs (satRNAs) are small subviral particles that are associated with the genomic RNA of a helper virus (HV). Their replication, encapsidation, and movement depend on the HV. In this paper, we performed a global analysis of the satRNAs associated with different isolates of tomato black ring virus (TBRV). We checked the presence of satRNAs in 42 samples infected with TBRV, performed recombination and genetic diversity analyses, and examined the selective pressure affecting the satRNAs population. We identified 18 satRNAs in total that differed in length and the presence of point mutations. Moreover, we observed a strong effect of selection operating upon the satRNA population. We also constructed infectious cDNA clones of satRNA and examined the viral load of different TBRV isolates in the presence and absence of satRNAs, as well as the accumulation of satRNA molecules on infected plants. Our data provide evidence that the presence of satRNAs significantly affects viral load; however, the magnitude of this effect differs among viral isolates and plant hosts. We also showed a positive correlation between the number of viral genomic RNAs (gRNAs) and satRNAs for two analysed TBRV isolates.

HLA-DQA1*05 Associates with Extensive Ulcerative Colitis at Diagnosis: An Observational Study in Children.

The human leukocyte antigen (HLA) allele group HLA-DQA1*05 predisposes to ulcerative colitis (UC) and is associated with the development of antibodies against infliximab in patients with inflammatory bowel disease (IBD). Therefore, we hypothesized that the presence of HLA-DQA1*05 correlates with characteristics of pediatric IBD. Within a multi-center cohort in Poland, the phenotype at diagnosis and worst flare was established and HLA-DQA1*05 status was assessed enabling genotype-phenotype analyses. HLA-DQA1*05 was present in 221 (55.1%) out of 401 children with IBD (UC n = 188, Crohn's disease n = 213). In UC, the presence of HLA-DQA1*05 was moderately associated with a large extent of colonic inflammation at diagnosis (E4 55% more frequent in HLA-DQA1*05-positive patients, p = 0.012). PUCAI at diagnosis (p = 0.078) and the time from UC diagnosis to the first administration of biologic treatment (p = 0.054) did not differ depending on HLA-DQA1*05 status. The number of days of hospitalization for exacerbation was analyzed in 98 patients for whom sufficient follow-up was available and did not differ depending on HLA-DQA1*05 carriership (p = 0.066). HLA-DQA1*05 carriers with CD were less likely to present with both stenosing and penetrating disease (B2B3, p = 0.048) and to have active disease proximal to the ligament of Treitz (L4a) at the worst flare (p = 0.046). Future research focusing on explaining and preventing anti-TNF immunogenicity should take into account that ADA may develop not only as an isolated reaction to anti-TNF exposure but also as a consequence of intrinsic differences in the early course of UC.

Genetic Variants of DMBT1 and SFTPD and Disease Severity in Paediatric Inflammatory Bowel Disease-A Polish Population-Based Study.

Deleted in malignant brain tumours 1 protein (DMBT1) and surfactant protein D (SFTPD) are antimicrobial peptides previously linked to inflammatory bowel disease (IBD) susceptibility. This study attempts to link the most potential IBD-associated polymorphisms in DMBT1 and SFTPD with the disease severity in children. A total of 406 IBD patients (Crohn's disease (CD) n = 214 and ulcerative colitis (UC) n = 192) were genotyped using hydrolysis probe assay. Clinical expression was described by disease activity scales, albumin and C-reactive protein levels, localisation and behaviour (Paris classification), systemic steroid, immunosuppressive, biological, and surgical treatment, number of exacerbation-caused hospitalisations, relapses and nutritional status. IBD patients with the risk genotype (AA) in DMBT1 rs2981804 had more frequent biological treatment (AA: vs. AG/GG; p = 0.012), concomitant diseases (AA vs. AG vs. GG; p = 0.015) and cutaneous manifestations (AA vs. AG/GG, p = 0.008). In UC, rs2981804 genotypes might be linked with albumin concentrations at diagnosis (AA vs. AG vs. GG; p = 0.009). In CD, DMBT1 rs2981745 was significantly associated with the number of severe relapses per year of disease (p = 0.020) and time-to-immunosuppression (p = 0.045). SFTPD was seemingly found to be associated with age at first immunosuppression in IBD (CC vs. CT vs. TT; p = 0.048). In conclusion, selected polymorphisms of DMBT1 and SFTPD might be associated with some disease severity measures in children with IBD. However, the magnitude of associations and their clinical relevance might be minor.

Apolipoprotein E variants correlate with the clinical presentation of paediatric inflammatory bowel disease: A cross-sectional study.

BACKGROUND: It has been suggested that apolipoprotein E (APOE) polymorphisms are associated with the risk of developing inflammatory bowel disease (IBD) and the early age of disease onset. However, there are no reports regarding the relationship with clinical characteristics and disease severity. AIM: To summarise that APOE polymorphisms are associated with the risk of developing IBD and the early age of disease onset. METHODS: In total, 406 patients aged 3-18 with IBD (192 had ulcerative colitis and 214 had Crohn's disease) were genotyped using the TaqMan hydrolysis probe assay. Clinical expression was described at diagnosis and the worst flare by disease activity scales, albumin and C-reactive protein levels, localisation and behaviour (Paris classification). Systemic steroid intake with the total number of courses, immunosuppressive, biological, and surgical treatment with the time and age of the first intervention were determined. The total number of exacerbation-caused hospitalisations, the number of days spent in hospital due to exacerbation, the number of relapses, and severe relapses were also estimated. RESULTS: Ulcerative colitis patients with the APOEε4 allele had lower C-reactive protein values at diagnosis (P = 0.0435) and the worst flare (P = 0.0013) compared to patients with the APOEε2 allele and genotype APOEε3/ε3. Crohn's disease patients with the APOEε2 allele scored lower on the Pediatric Crohn's Disease Activity Index at diagnosis (P = 0.0204). IBD patients with APOEε2 allele spent fewer days in the hospital due to relapse (P = 0.0440). CONCLUSION: APOE polymorphisms are associated with the risk of developing IBD and the clinical expression of IBD. However, the clinical relevance of the differences identified is rather modest.

Leukocyte Telomere Length Is Not Reduced in Children and Adults with Cystic Fibrosis but Associates with Clinical Characteristics-A Cross-Sectional Study.

We hypothezied that telomere length is considerably altered in cystic fibrosis (CF) patients compared to healthy subjects (HS), and that leukocyte telomere length variation reflects the severity of CF. Relative telomere length (RTL) was assessed by qPCR in 70 children aged 5-10 (34 CF; 36 HS) and 114 adults aged 18-45 (53 CF; 61 HS). Telomere length was similar in CF and HS (median (interquartile range): 0.799 (0.686-0.950) vs. 0.831 (0.707-0.986); p = 0.5283) both in children and adults. In adults, women had longer telomeres than men (0.805 (0.715-0.931) vs. 0.703 (0.574-0.790); p = 0.0002). Patients treated with inhaled corticosteroids had a shorter RTL compared to those without steroid therapy (0.765 (0.664-0.910) vs. 0.943 (0.813-1.191); p = 0.0007) and this finding remained significant after adjusting for gender, age, BMI, and child/adult status (p = 0.0003). Shorter telomeres were independently associated with the presence of comorbidities (0.763 (0.643-0.905) vs. 0.950 (0.783-1.130); p = 0.0006) and antibiotic treatment at the moment of blood sampling (0.762 (0.648-0.908) vs. 0.832 (0.748-1.129); p = 0.0172). RTL correlated with number of multiple-day hospitalizations (rho = -0.251; p = 0.0239), as well as number of hospitalization days (rho = -0.279; p = 0.0113). Leukocyte RTL in children and adults with CF was not shorter than in healthy controls, and did not seem to have any potential as a predictor of CF survival. However, it inversely associated with the investigated clinical characteristics.

Subjective Psychophysical Experiences in the Course of Inflammatory Bowel Disease-A Comparative Analysis Based on the Polish Pediatric Crohn's and Colitis Cohort (POCOCO).

No gold standard is available to evaluate subjective psychophysical experiences in pediatric inflammatory bowel disease (IBD). We aimed to assess pain, anxiety, and limitations in social activities at diagnosis and the worst flare of the disease in relation to clinical expression, treatment and IBD severity. A total of 376 children completed the survey (Crohn's disease (CD) n = 196; ulcerative colitis (UC) n = 180). The questionnaire included 12 questions regarding pain, anxiety, and social activity, all assessed at recruitment and retrospectively at diagnosis and worst flare using a numeric rating scale. Patients that had ever been treated with systemic glucocorticosteroids scored higher in pain (p < 0.001), anxiety (p = 0.015), and social activity domains (p < 0.016) at worst flare, and the answers correlated with the number of steroid courses (p < 0.0392). The perception of social activity limitations also correlated independently with the number of immunosuppressants (p < 0.0433) and biological agents (p < 0.0494). There was no difference in retrospective perception of pain, anxiety and social activity limitations between CD and UC patients at diagnosis and the worst flare. The level of limitations in social activity correlated with hospitalisations due to relapse, days spent in the hospital, number of relapses, and severe relapses with the strongest association of rho = 0.39 (p = 0.0004). Subjective and retrospective perception of pain, anxiety, and limitations in social activity differs depending on therapy, correlates with treatment modalities, and severity measures such as hospitalisations.

Insolation and Disease Severity in Paediatric Inflammatory Bowel Disease-A Multi-Centre Cross-Sectional Study.

This study was to investigate whether the clinical course of inflammatory bowel disease (IBD) in a Polish paediatric cohort fits a seasonal pattern and depends on insolation. Two hundred and fourteen patients diagnosed with Crohn's disease (CD) and 192 with ulcerative colitis (UC) aged from 3 to 18 years, were recruited in seven centres of similar latitude. The seasons were defined as winter (December-February), spring (March-May), summer (June-August), autumn (September-November). The year was also divided depending on insolation threshold (3.0 kWh/m2/day). Patients diagnosed with IBD when the isolation was >3 kWh/m2/day had poorer nutritional status than those diagnosed while insolation was below threshold (lower standardised BMI at diagnosis (-0.81 ([-1.34]-[-0.03]) vs. -0.52 ([-1.15]-0.15); p = 0.0320) and worst flare (-0.93 ([-1.37]-[-0.05]) vs. -0.66 ([-1.23]-0.17); p = 0.0344), with the need for more frequent biological treatment (45.5% vs. 32.7%, p = 0.0100). Patients diagnosed in winter were significantly younger at diagnosis (11.4 vs. 13.0; padj = 0.0180) and first immunosuppressive treatment (11.3 vs. 13.3; padj = 0.0109) than those diagnosed in other seasons. CD patients diagnosed in months with higher insolation spent more days in hospital than those diagnosed in months with lower insolation [4.6 (1.8-11.8) vs. 2.9 (1.3-6.2); p = 0.0482]. CD patients diagnosed in summer had significantly more concomitant diseases. In patients with CD, the occurrence of the worst flare was more frequent in autumn. Furthermore, the season of birth was associated with Pediatric Crohn's Disease Activity Index at worst flare and earlier surgery. In conclusion, several clinical parameters are associated with insolation, the season of diagnosis and season of birth in the clinical course of Crohn's disease.

The role of CaO/SiO2 ratio and P2O5 content in gel-derived bioactive glass-polymer composites in the modulation of their bioactivity and osteoinductivity in human BMSCs.

We obtained a range of PLGA-based composites containing sol-gel bioactive glasses (SBG) from the SiO2-CaO and SiO2-CaO-P2O5 systems. Eight SBGs with different CaO/SiO2 ratios with and without P2O5 were incorporated at 50% w/w to PLGA matrix and structured into thin films suitable for cell culture. The SBG/PLGA composites were examined for their bioactivity in simulated body fluid (SBF), ion release profile in culture media with and without cells, and osteoinductivity in standard human bone marrow stromal cell (hBMSC) cultures without osteogenic growth factors. Our results indicate different surface activity of composites depending on the presence/absence of P2O5 in SBG composition. Furthermore, ion release profile to culture medium differed depending on the presence/absence of cells. Direct culture of hBMSC on the SiO2-CaO/PLGA composite films resulted in elevated Runx-2 mRNA, opposite to low Runx-2 mRNA levels on SiO2-CaO-P2O5/PLGA films. All studied composites increased Osx mRNA levels. Whereas some of SiO2-CaO/PLGA composites did not elevate BMP-2 and -6 proteins in hBMSC cultures, high levels of these BMPs were present in all cultures on SiO2-CaO-P2O5/PLGA composites. All composites induced BMP-related Tak1 signalling, whereas Smad1 signalling was restricted mostly to composites containing three-component SBGs. ALP activity of hBMSC and BMP-related luciferase activity of mouse BRITE cells differed depending on whether the cells were stimulated with culture medium conditioned with SBG/PLGA composites or the cells were directly cultured on the composite surfaces. Altogether, beyond bioactivity and osteoinductivity of SBG/PLGA composites, our studies show key differences in the biological response to both the bioactive material dissolution products and upon direct cell-material contacts.

Multifunctional Polymer Composites Produced by Melt-Blown Technique to Use in Filtering Respiratory Protective Devices.

In this work, a multifunctional polymer composite is made using melt-blowing technology from polypropylene (88 wt.%) and poly (ethylene terephthalate) (12 wt.%) with the addition of functional modifiers, that is, 3 g of a superabsorbent polymer and 5 g of a biocidal agent (Biohaloysite). The use of modifiers is aimed at obtaining adequate comfort when using the target respiratory protection equipment (RPE) in terms of microclimate in the breathing zone and protection against harmful aerosols including bioaerosols. The developed production method is innovative in that the two powdered modifiers are simultaneously applied in the stream of elementary polymeric fibers by two independent injection systems. Aerosols of the modifiers are supplied via a specially designed channel in the central segment of the die assembly, reducing the amount of materials used in the production process and saving energy. The results show that the proposed method of incorporating additives into the fiber structure did not adversely affect the protective and functional properties of the resulting filtration nonwovens. The produced nonwoven composites are characterized by SEM, FTIR, and differential scanning calorimetry (DSC). Given their high filtration efficiency at 5%, satisfactory airflow resistance (~200 Pa), very good antimicrobial activity, and excellent water absorption capacity, the obtained multifunctional nonwoven composites may be successfully used in filtering respiratory protective devices.

Sulfatide decreases the resistance to stress-induced apoptosis and increases P-selectin-mediated adhesion: a two-edged sword in breast cancer progression.

BACKGROUND: We have previously shown that galactosylceramide (GalCer) affects the tumourigenic and metastatic properties of breast cancer cells by acting as an anti-apoptotic molecule. Since GalCer is a precursor molecule in the synthesis of sulfatides, the present study was aimed to define the role of sulfatides in apoptosis and breast cancer progression. METHODS: Expression of GAL3ST1 in breast cancer cell lines and breast cancer tissue specimens was analysed using real-time PCR, western blotting and immunohistochemistry analysis. The amount of sulfatide, GalCer and ceramide was analysed by thin-layer chromatography binding assay and by the modified hydrophilic interaction liquid chromatography coupled with electrospray mass spectrometry methodology. The tumourigenicity of cancer cells was analysed by an in-vivo tumour growth assay. Apoptotic cells were detected based on caspase-3 activation and the TUNEL assay. The interaction of breast cancer cells with P-selectin or E-selectin was analysed using the flow adhesion assay. The ability of sulfatide-expressing cells to activate and aggregate platelets was studied using the flow-cytometry-based aggregation assay. RESULTS: Using two models of breast cancer, T47D cells with blocked synthesis of sulfatide and MDA-MB-231 cells with neosynthesis of this glycosphingolipid, we showed that high sulfatide levels resulted in increased sensitivity of cancer cells to apoptosis induced by hypoxia and doxorubicin in vitro, and decreased their tumourigenicity after transplantation into athymic nu/nu mice. Accordingly, a clinical study on GAL3ST1 expression in invasive ductal carcinoma revealed that its elevated level is associated with better prognosis. Using MDA-MB-231 cells with neosynthesis of sulfatide we also showed that sulfatide is responsible for adhesion of breast cancer cells to P-selectin-expressing cells, including platelets. Sulfatide also acted as an activating molecule, increasing the expression of P-selectin. CONCLUSIONS: This study demonstrates that increased synthesis of sulfatide sensitises cancer cells to microenvironmental stress factors such as hypoxia and anticancer drugs such as doxorubicin. However, sulfatide is probably not directly involved in apoptotic cascades, because its increased synthesis by GAL3ST1 decreased the amounts of its precursor, GalCer, a known anti-apoptotic molecule. On the other hand, our data support the view that sulfatides are malignancy-related adhesive molecules involved in activating and binding P-selectin-expressing platelets to breast cancer cells.

Comparison of Microvessel Density Using Nestin and CD34 in Colorectal Cancer.

BACKGROUND/AIM: The aim of the present study was to compare the prognostic value of nestin-positive microvessel density (nestin+ MVD) to the CD34-positive (CD34+) MVD in patients with colorectal cancer. MATERIALS AND METHODS: A total of 125 cases of colorectal cancer were used for the study. Immunohistochemical reactions were performed on paraffin sections using anti-nestin and anti-CD34. The average MVD was determined for both markers and was referenced to patients' clinicopathological data. RESULTS: A strong positive correlation was found between the MVDs for both analyzed antigens. Contrary to CD34+ MVD, a positive trend was observed between high nestin+ MVD and lymph node invasion. Moreover, significantly higher nestin+ MVD was noted in patients with more advanced clinical stage than in those with early-stage cancer. No such correlation was found for CD34 antigen. Additionally, a possible trend between the higher nestin+ MVD and shorter event-free survival was noted, whereas no such relation was observed for CD34+ MVD. CONCLUSION: The results suggest that nestin might be a reliable marker for angiogenesis evaluation in patients with colorectal cancer. In contrast to CD34, the expression of nestin in blood vessels seems to be a more sensitive marker of cancer progression.

Implications of nestin in breast cancer pathogenesis (Review).

The aim of the present review was to summarize the current knowledge of the involvement of nestin in breast cancer (BC) pathogenesis. Nestin is a member of the class VI family of intermediate filament proteins, originally identified as a marker of neural stem cells and subsequently demonstrated to be expressed in BC and other cancer types. In normal breast tissue, nestin is expressed in the basal/myoepithelial cells of the mammary gland. In BC, nestin identifies basal-like tumours and predicts aggressive behaviour and poor prognosis. Nestin expression has also been detected in BC stem cells and newly-formed tumour vessels, being a factor in promoting invasion and metastasis. The present review provides an up-to-date overview of the involvement of nestin in processes facilitating BC pathogenesis and progression.

The Role of CHI3L1 Expression in Angiogenesis in Invasive Ductal Breast Carcinoma.

BACKGROUND/AIM: An increased level of chitinase 3 like 1 protein (CHI3L1) expression is observed in patients with cancer and may have potential prognostic value. The aim of this study was to evaluate the role of CHI3L1 in angiogenesis in invasive ductal breast carcinoma (IDC) (n=110). MATERIALS AND METHODS: Immunohistochemistry was used to assess the expression of CHI3L1, CD31, CD34, vascular endothelial growth factor (VEGFA, VEGFC and VEGFD). Real-time polymerase chain reaction and western blot were used to determine the level of CHI3L1 mRNA and protein. RESULTS: Immunohistochemistry demonstrated positive correlation between CHI3L1 expression and angiogenesis markers: CD31 (r=0.34, p=0.0003), CD34 (r=0.24, p=0.012), VEGFD (r=0.24, p=0.013). Higher CHI3L1 expression in estrogen receptor-negative (p=0.041) and progesterone receptor-negative (p=0.014) cancer was observed. Higher CHI3L1 expression was reported in cancer tissues in comparison to non-malignant breast lesions. CONCLUSION: These results suggest a potential role of CHI3L1 in angiogenesis in IDC and may suggest its involvement in cancer progression.

The expression of IL10RA in colorectal cancer and its correlation with the proliferation index and the clinical stage of the disease.

INTRODUCTION: Despite the widely described role of IL10 in immune response regulation during carcinogenesis, there is no established model describing the role of its receptor. The aim of this study is to elucidate the relationship between the subunit alpha of IL10 receptor (IL10RA) in the pathogenesis of colorectal cancer (CRC). METHODS: The study was conducted on archived paraffin blocks of 125 CRC patients, from which tissue microarrays (TMA) were made. These were subsequently used for immunohistochemistry to assess the expression of IL10RA, IL10, phosphorylated STAT3 (pSTAT3) and the Ki67 proliferation index. The intensity of both reactions was assessed by independent researchers using two approaches: digital image analysis and the Remmele and Stegner score (IRS). To assess the possible correlations between the two investigated markers and the clinical stage of CRC, the Pearson correlation coefficient was calculated. The expression of aforementioned proteins was assessed in tumor samples, healthy surgical margins and healthy control samples, obtained from cadavers during autopsy from the Department of Forensic Medicine. Statistical analysis was conducted using Statistica ver. 13.05 software. RESULTS: The final analysis included 105 CRC patients with complete clinical and pathological data, for whom the expressions of IL10RA, IL10, pSTAT3 and Ki67 were assessed using two independent methods. There was a positive correlation between the IL10RA expression and Ki-67 proliferation index (R = 0.63, p < 0.001) and a negative correlation between the IL10RA expression and the clinical stage of CRC (R = -0.21, p = 0.022). IL10RA correlated positively with pSTAT3 and IL10 in neoplastic tissue and tumor margin (with p < 0.01 for all correlations). We also observed a significantly higher expression of IL10RA in healthy surgical margins when compared to the actual tumor (p = 0.023, the paired t-test). The expression of IL10 was significantly higher in tumors than in healthy intestinal endothelium from control group. CONCLUSIONS: The correlations between the expression of IL10RA and the proliferation index or the clinical stage of CRC seem to confirm the importance of IL10RA in the pathogenesis of CRC. The higher expression of IL10RA in healthy surgical margins than in the tumor itself may suggest that IL10RA plays a role in regulating immune response to the neoplasm.

Defective RNA particles derived from Tomato black ring virus genome interfere with the replication of parental virus.

Tomato black ring virus (TBRV) is the only member of the Nepovirus genus that is known to form defective RNA particles (D RNAs) during replication. Here, de novo generation of D RNAs was observed during prolonged passages of TBRV isolates originated from Solanum lycopersicum and Lactuca sativa in Chenopodium quinoa plants. D RNAs of about 500 nt derived by a single deletion in the RNA1 molecule and contained a portion of the 5' untranslated region and viral replicase, and almost the entire 3' non-coding region. Short regions of sequence complementarity were found at the 5' and 3' junction borders, which can facilitate formation of the D RNAs. Moreover, in this study we analyzed the effects of D RNAs on TBRV replication and symptoms development of infected plants. C. quinoa, S. lycopersicum, Nicotiana tabacum, and L. sativa were infected with the original TBRV isolates (TBRV-D RNA) and those containing additional D RNA particles (TBRV + D RNA). The viral accumulation in particular hosts was measured up to 28 days post inoculation by RT-qPCR. Statistical analyses revealed that D RNAs interfere with TBRV replication and thus should be referred to as defective interfering particles. The magnitude of the interference effect depends on the interplay between TBRV isolate and host species.