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Nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome activation plays an important role in chronic obstructive pulmonary disease (COPD) pathogenesis and might be involved in ongoing chronic inflammation. This study aimed to determine interleukin-1beta (IL-1β) plasma concentration as well as IL1B , NLRP3 and caspase-1 ( CASP1 ) gene expression in the Croatian COPD patients. 109 patients with stable COPD and age- and sex-matched 95 controls were included in the study. Plasma IL-1β concentration was measured by Luminex technology, and gene expression analysis was performed using TaqMan assays. It was shown that COPD patients had increased concentration of IL-1β and enhanced gene expression of IL1B , NLRP3 and CASP1 compared to controls. There was no difference in IL-1β or IL1B , NLRP3 and CASP1 in patients with COPD regarding airflow obstruction severity and smoking history. Finally, the diagnostic potential of the determined parameters was evaluated, and it was found that IL-1β correctly classified 89% of cases in the combination with common inflammatory biomarkers, white blood cell count and fibrinogen, showing a potential in COPD prediction. In conclusion, up-regulation of IL1B , NLRP3 , CASP1 and increased IL-1β concentration suggest the activation of NLRP3 inflammasome in the systemic compartment of patients with stable COPD.

Colorectal cancer (CRC) is the third most common cancer worldwide. The high mortality from CRC is mainly related to metastasis affecting distant organs and their function. Dissemination of tumor cells from the primary tumor and hematogeneous spread are considered crucial in the formation of tumor metastases. The analysis of circulating tumor cells (CTCs) and CTC clusters in the blood can be used for the early detection of invasive cancer. Moreover, CTCs have a prognostic significance in the monitoring of a malignant disease or the response to chemotherapy. This work presents an overview of the research conducted on CTCs with the aim of finding suitable detection systems and assessing the possibility of clinical applications in patients with CRC.

Specific markers for colorectal cancer (CRC), preceded by colorectal adenoma (pre-CRC), are lacking. This study aimed to investigate whether microRNAs (miR-19a-3p, miR-92a-3p, miR-193a-3p, and miR-210-3p) from tissues and exosomes are potential CRC biomarkers and compare them to existing biomarkers, namely carcinoembryonic antigen (CEA) and carbohydrate antigen (CA) 19-9. MiRNA was isolated in the samples of 52 CRC and 76 pre-CRC patients. Expression levels were analyzed by RT-qPCR. When comparing pre-CRC and CRC tissue expression levels, only miR-193a-3p showed statistically significant result (p < 0.0001). When comparing the tissues and exosomes of CRC samples, a statistically significant difference was found for miR-193a-3p (p < 0.0001), miR-19a-3p (p < 0.0001), miR-92a-3p (p = 0.0212), and miR-210-3p (p < 0.0001). A receiver-operating characteristic (ROC) curve and area under the ROC curve (AUC) were used to evaluate the diagnostic value of CEA, CA 19-9, and miRNAs. CEA and CA 19-9 had good diagnostic values (AUCs of 0.798 and 0.668). The diagnostic value only of miR-193a-3p was highlighted (AUC = 0.725). The final logistic regression model, in which we put a combination of CEA concentration and the miR-193a-3p expression level in tissues, showed that using these two markers can distinguish CRC and pre-CRC in 71.3% of cases (AUC = 0.823). MiR-193a-3p from tissues could be a potential CRC biomarker.

Drug-specific therapeutic approaches for colorectal cancer (CRC) have contributed to significant improvements in patient health. Nevertheless, there is still a great need to improve the personalization of treatments based on genetic and epigenetic tumor profiles to maximize the quality and efficacy while limiting cytotoxicity. Currently, CEA and CA 19-9 are the only validated blood biomarkers in clinical practice. For this reason, laboratories are trying to identify new specific prognostics and, more importantly, predictive biomarkers for CRC patient profiling. Thus, the unique landscape of personalized biomarker data should have a clinical impact on CRC treatment strategies and molecular genetic screening tests should become the standard method for diagnosing CRC. This review concentrates on recent molecular testing in CRC and discusses the potential modifications in CRC assay methodology with the upcoming clinical application of novel genomic approaches. While mechanisms for analyzing circulating tumor DNA have been proven too inaccurate, detecting and analyzing circulating tumor cells and protein analysis of exosomes represent more promising options. Blood liquid biopsy offers good prospects for the future if the results align with pathologists’ tissue analyses. Overall, early detection, accurate diagnosis and treatment monitoring for CRC with specific markers and targeted molecular testing may benefit many patients.

Heat shock protein 70 (Hsp70) engages Toll-like receptors (TLR) 2 and 4 when found in the extracellular compartment and contributes to inflammation in chronic obstructive pulmonary disease (COPD). Since there is growing evidence for the genetic risk factors for COPD, the gene expression of HSP70, TLR2 and TLR4 was determined, as well as the association between HSP70, TLR2 and TLR4 single nucleotide polymorphisms, (SNPs) and COPD. The gene expression was assessed in peripheral blood cells of 137 COPD patients and 95 controls by a quantitative polymerase chain reaction (qPCR), while a total of nine SNPs were genotyped by TaqMan allelic discrimination real-time PCR. HSP70 and TLR2 gene expression was increased in COPD patients compared to the controls, regardless of the disease severity and smoking status of participants. The rs6457452 SNP of HSP70 was associated with COPD, indicating the protective role of the T allele (OR = 0.46, 95% CI = 0.24–0.89, p = 0.022). Furthermore, COPD C/T heterozygotes showed a decreased HSP70 mRNA level compared to COPD C/C homozygotes. In conclusion, HSP70 and TLR2 may have a role in the pathogenesis of COPD, and the HSP70 rs6457452 variant might influence the genetic susceptibility to COPD in the Croatian population.

Liquid biopsy has an underexplored diagnostic potential in colorectal cancer (CRC). Sufficient quantity and quality of its elements (circulating cell-free DNA (ccfDNA), exosomes and exosomal RNA) are essential for accurate results. The present study aims to establish the optimal protocol for handling liquid biopsy samples. Samples were obtained by collecting peripheral blood from colorectal adenoma patients in CellSave tubes. Plasma was separated within six hours using differential centrifugation and aliquots stored at – 20/– 80 °C until further processing. Three methods for isolation of ccfDNA, and two combinations of kits for isolation of exosomes and exosomal RNA were tested. The quality and quantity of ccfDNA isolates were evaluated. Exosomes were characterised by determining size, concentration, and total and specific protein content. Expression of chosen microRNAs, miR-19a-3p and miR-92-3p, which have been implicated in CRC progression, were determined. The vacuum-column-based kit showed the highest quantities of isolated ccfDNA (P-value < 0.001). Kits for exosome isolation significantly differed in size (P-value = 0.016), concentration (P-value = 0.016) and protein content (P-value = 0.016). There was no significant difference in expressions of miR-19a-3p (P-value = 0.219) and miR-92a-3p (P-value = 0.094) between the two isolation kits. The new, adapted protocol described, enables simultaneous analysis of multiple elements when investigating potential biomarkers of CRC.

Background High resolution melting (HRM) analysis is one of the newer, reliable, and sensitive genotyping techniques, which offers considerable time and cost savings. P‐selectin is an adhesion molecule that has a role in the initial phases of leukocyte adhesion to stimulated platelets and endothelial cells in inflammation. Multiple polymorphisms in P‐selectin gene (SELP) that affect the protein sequence have been described. The aim of this study was to design, optimize, and validate a simple and rapid in‐house HRM‐based method for genotyping the NM_003005.3:c.992G>A (c.992G>A), NM_003005.3:c.1918G>T (c.1918G>T), and NM_003005.3:c.2266A>C (c.2266A>C) SELP polymorphisms. Methods Initial genotyping of three SELP polymorphisms was performed by applying polymerase chain reaction (PCR) with sequence‐specific primers (SSP), which was used as a reference method for determination of analytical sensitivity. PCR‐HRM was performed with primers for c.2266A>C reported in the literature. Primers for the remaining two polymorphisms were designed using Primer‐BLAST. Precision testing was performed using three samples with different genotypes. For accuracy, analytical sensitivity and specificity testing, 20 wild type, 10 heterozygous, and 10 homozygous samples were chosen per polymorphism. Results were expressed as percentage of concordance with the acceptability criterion ≥95%. Results Agreement of results was 100% for all validation parameters except for analytical sensitivity for c.1918G>T and c.2266A>C, with agreement of 90%. Repeated analysis using both methods revealed an error in initial genotyping and correct genotyping by PCR‐HRM, which was confirmed by Sanger sequencing. Conclusion The validation confirmed PCR‐HRM as a precise, accurate, and specific method for genotyping the c.992G>A, c.1918G>T, and c.2266A>C SELP polymorphisms.

Lockdowns implemented to address the COVID-19 pandemic have disrupted human mobility flows around the globe to an unprecedented extent and with economic consequences which are unevenly distributed across territories, firms and individuals. Here we study socioeconomic determinants of mobility disruption during both the lockdown and the recovery phases in Italy. For this purpose, we analyze a massive data set on Italian mobility from February to October 2020 and we combine it with detailed data on pre-existing local socioeconomic features of Italian administrative units. Using a set of unsupervised and supervised learning techniques, we reliably show that the least and the most affected areas persistently belong to two different clusters. Notably, the former cluster features significantly higher income per capita and lower income inequality than the latter. This distinction persists once the lockdown is lifted. The least affected areas display a swift (V-shaped) recovery in mobility patterns, while poorer, most affected areas experience a much slower (U-shaped) recovery: as of October 2020, their mobility was still significantly lower than pre-lockdown levels. These results are then detailed and confirmed with a quantile regression analysis. Our findings show that economic segregation has, thus, strengthened during the pandemic.

SARS-CoV-2 viremia has been found to be a potential prognostic factor in patients hospitalized for COVID-19. We aimed to assess the association between SARS-CoV-2 viremia and mortality in COVID-19 hospitalized patients during different epidemic periods. A prospective COVID-19 registry was queried to extract all COVID-19 patients with an available SARS-CoV-2 viremia performed at hospital admission between March 2020 and January 2022. SARS-CoV-2 viremia was assessed by means of GeneFinderTM COVID-19 Plus RealAmp Kit assay and SARS-CoV-2 ELITe MGB® Kit using <45 cycle threshold to define positivity. Uni and multivariable logistic regression model were built to assess the association between SARS-CoV-2 positive viremia and death. Four hundred and forty-five out of 2,822 COVID-19 patients had an available SARS-CoV-2 viremia, prevalently males (64.9%) with a median age of 65 years (IQR 55-75). Patients with a positive SARS-CoV-2 viremia (86/445; 19.3%) more frequently presented with a severe or critical disease (67.4% vs 57.1%) when compared to those with a negative SARS-CoV-2 viremia. Deceased subjects (88/445; 19.8%) were older [75 (IQR 68-82) vs 63 (IQR 54-72)] and showed more frequently a detectable SARS-CoV-2 viremia at admission (60.2% vs 22.7%) when compared to survivors. In univariable analysis a positive SARS-CoV-2 viremia was associated with a higher odd of death [OR 5.16 (95% CI 3.15-8.45)] which was confirmed in the multivariable analysis adjusted for age, biological sex and, disease severity [AOR 6.48 (95% CI 4.05-10.45)]. The association between positive SARS-CoV-2 viremia and death was consistent in the period 1 February 2021-31 January 2022 [AOR 5.86 (95% CI 3.43-10.16)] and in subgroup analysis according to disease severity: mild/moderate [AOR 6.45 (95% CI 2.84-15.17)] and severe/critical COVID-19 patients [AOR 6.98 (95% CI 3.68-13.66)]. SARS-CoV-2 viremia resulted associated to COVID-19 mortality and should be considered in the initial assessment of COVID-19 hospitalized patients.