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Background: The colorectal adenoma undergoes neoplastic progression via the normal epithelium–adenoma–adenocarcinoma sequence as reported in the Vogelgram. The hazard of developing a tumor is deeply associated with the number and size of adenomas and their subtype. Adenomatous polyps are histologically categorized as follows: approximately 80–90% are tubular, 5–15% are villous, and 5–10% are tubular/villous. Given the higher risk of a malignant transformation observed in tubular/villous adenomas, patients diagnosed with adenomatous polyposis are at an improved risk of developing CRC. The Wnt/β-catenin pathway plays a key role in the onset of colorectal adenoma; in particular, intestinal cells first acquire loss-of-function mutations in the APC gene that induce the formation of adenomas. Methods: Wnt/β-catenin pathway APC, Wnt3a, Wnt5a, LEF1, and BCL9 genes and protein expression analyses were conducted by qRT-PCR and western blot in 68 colonic samples (polyps and adjacent mucosa) from 41 patients, of which 17 were affected by FAP. Ten normal colonic mucosal samples were collected from 10 healthy donors. Results: In this study, both the APC gene and protein were less expressed in the colon tumor compared to the adjacent colonic mucosa. Conversely, the activated β-catenin was more expressed in polyps than in the adjacent mucosa. All results confirmed the literature data on carcinomas. A statistically significant correlation between Wnt3a and BCL9 both in polyps and in the adjacent mucosa underlines that the canonical Wnt pathway is activated in early colon carcinogenesis and that the adjacent mucosa is already altered. Conclusion: This is the first study analyzing the difference in expression of the Wnt/β-catenin pathway in human colorectal adenomas. Understanding the progression from adenomas to colorectal carcinomas is essential for the development of new therapeutic strategies and improving clinical outcomes with the use of APC and β-catenin as biomarkers.

In the tumor microenvironment (TME), ROS production affects survival, progression, and therapy resistance in colorectal cancer (CRC). H2O2-mediated oxidative stress can modulate Wnt/β-catenin signaling and metabolic reprogramming of the TME. Currently, it is unclear how mild/moderate oxidative stress (eustress) modulates Wnt/β-catenin/APC and JNK signaling relationships in primary and metastatic CRC cells. In this study, we determined the effects of the H2O2 concentration inducing eustress on isogenic SW480 and SW620 cells, also in combination with JNK inhibition. We assessed cell viability, mitochondrial respiration, glycolysis, and Wnt/β-catenin/APC/JNK gene and protein expression. Primary CRC cells were more sensitive to H2O2 eustress combined with JNK inhibition, showing a reduction in viability compared to metastatic cells. JNK inhibition under eustress reduced both glycolytic and respiratory capacity in SW620 cells, indicating a greater capacity to adapt to TME. In primary CRC cells, H2O2 alone significantly increased APC, LEF1, LRP6, cMYC and IL8 gene expression, whereas in metastatic CRC cells, this effect occurred after JNK inhibition. In metastatic but not in primary tumor cells, eustress and inhibition of JNK reduced APC, β-catenin, and pJNK protein. The results showed differential cross-regulation of Wnt/JNK in primary and metastatic tumor cells under environmental eustress conditions. Further studies would be useful to validate these findings and explore their therapeutic potential.

This study aims to test a photodynamic protocol based on a gel containing aminolevulinic acid followed by red-LED (ALAD-PDT) irradiation on human gingival fibroblasts (hGFs) and osteoblasts (hOBs) cultured on a porcine acellular dermal matrix membrane (PADMM). In the previous literature, ALAD-PDT showed solid antibacterial activity and proliferative induction on HGFs cultured on plates and HOBs cultured on a cortical lamina. PADMMs are used in dentistry and periodontology to treat gingival recessions and to increase the tissue thickness in the case of a thin biotype without the risks or postoperative discomfort associated with connective tissue grafts. However, one of the possible complications in this type of surgery is represented by bacterial invasion and membrane exposition during the healing period. We hypothesized that the addition of ALAD-PDT to PADMMs could enhance more rapid healing and decrease the risks connected with bacterial invasion. In periodontal surgery, PADMMs are inserted after a full-thickness flap elevation between the bone and the flap. Consequently, all procedures were performed in parallel on hOBs and hGFs obtained by dental patients. The group control (CTRL) was represented by the unexposed cells cultured on the membranes, group LED (PDT) were the cells subjected to 7 min of red LED irradiation, and ALAD-PDT were the cells subjected to 45 min of ALAD incubation and then to 7 min of red LED irradiation. After treatments, all groups were analyzed for MTT assay and subjected to histological examination at 3 and 7 days and to the SEM observations at 3, 7, and 14 days. Different bone mineralization assays were performed to quantify the effects of ALAD-PDT on hOBs: ALP activity, ALP gene expression, osteocalcin, and alizarin red. The effects of ALAD-PDT on hGFs were evaluated by quantifying collagen 1, fibronectin, and MMP-8. Results showed that ALAD-PDT promoted cellular induction, forming a dense cellular network on hOBs and hGFs, and the assays performed showed statistically significantly higher values for ALAD-PDT with respect to LED alone and CTRLs. In conclusion, ALAD-PDT could represent a promising aid for enhancing the healing of gingival tissues after PADMM applications.

Interaction between infectious agents and liver tissue, as well as repeated and extreme biological events beyond adaptive capacities, may result in pathological conditions predisposing people to development of primary liver cancers (PLCs). In adults, PLCs mainly comprise hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA). Various infectious agents in the hepatic microenvironment can destabilize normal liver cell functions by modulating the Wnt/β-catenin pathway components. Among them, hepatotropic viruses B, C, and D are involved in Wnt/β-catenin signaling dysregulation. Other microbial agents, including oncogenic viruses such as Epstein–Barr virus (EBV) and human papilloma virus (HPV), bacteria, e.g., Mycoplasma hyorhinis and Salmonella Typhi, the protozoan parasite Toxoplasma gondii, the fungus Aspergillus flavus, and liver flukes such as Clonorchissinensis or Opisthorchis viverrini, may induce malignant transformation in hepatocytes or in target cells of the biliary tract through aberrant Wnt signaling activation. This review focuses on new insights into infectious agents implicated in the deregulation of Wnt signaling and PLC development. Since the Wnt/β-catenin pathway is a driver of cancer following viral and bacterial infections, molecules inhibiting the complex axis of Wnt signaling could represent novel therapeutic approaches in PLC treatment.

Gastric cancer is worldwide the fifth and third cancer for incidence and mortality, respectively. Stomach wall is daily exposed to oxidative stress and BER system has a key role in the defense from oxidation-induced DNA damage, whilst ErbB receptors have important roles in the pathogenesis of cancer. We used AGS cells as an aggressive gastric carcinoma cell model, treated with H2O2 alone or combined with ErbB signaling pathway inhibitors, to evaluate the effects of oxidative stress in gastric cancer, focusing on the modulation of ErbB signaling pathways and their eventual cross-talk with BER system. We showed that treatment with H2O2 combined with PI3K/AKT and MEK inhibitors influenced cell morphology and resulted in a reduction of cancer cell viability. Migration ability was reduced after H2O2 treatment alone or combined with MEK inhibitor and after PI3K/AKT inhibitor alone. Western blotting analysis showed that oxidative stress stimulated EGFR pathway favoring the MAPKs activation at the expense of PI3K/AKT pathway. Gene expression analysis by RT-qPCR showed ErbB2 and OGG1 increase under oxidative stress conditions. Therefore, we suggest that in AGS cells a pro-oxidant treatment can reduce gastric cancer cell growth and migration via a different modulation of PI3K and MAPKs pathways. Moreover, the observed ErbB2 and OGG1 induction is a cellular response to protect the cells from H2O2-induced cell death. In conclusion, to tailor specific combinations of therapies and to decide which strategy to use, administration of a chemotherapy that increases intracellular ROS to toxic levels, might not only be dependent on the tumor type, but also on the molecular targeting therapy used.

Thyroid diseases have a complex and multifactorial aetiology. Despite the numerous studies on the signals referable to the malignant transition, the molecular mechanisms concerning the role of oxidative stress remain elusive. Based on its strong oxidative power, H2O2 could be responsible for the high level of oxidative DNA damage observed in cancerous thyroid tissue and hyperactivation of mitogen-activated protein kinase (MAPK) and PI3K/Akt, which mediate ErbB signaling. Increased levels of 8-oxoG DNA adducts have been detected in the early stages of thyroid cancer. These DNA lesions are efficiently recognized and removed by the base excision repair (BER) pathway initiated by 8-oxoG glycosylase1 (OGG1). This study investigated the relationships between the EGFR and OGG1-BER pathways and their mutual regulation following oxidative stress stimulus by H2O2 in human thyrocytes. We clarified the modulation of ErbB receptors and their downstream pathways (PI3K/Akt and MAPK/ERK) under oxidative stress (from H2O2) at the level of gene and protein expression, according to the mechanism defined in a human non-pathological cell system, Nthy-ori 3-1. Later, on the basis of the results obtained by gene expression cluster analysis in normal cells, we assessed the dysregulation of the relationships in a model of papillary thyroid cancer with RET/PTC rearrangement (TPC-1). Our observations demonstrated that a H2O2 stress may induce a physiological cross-regulation between ErbB and OGG1-BER pathways in normal thyroid cells (while this is dysregulated in the TPC-1 cells). Gene expression data also delineated that MUTYH gene could play a physiological role in crosstalk between ErbB and BER pathways and this function is instead lost in cancer cells. Overall, our data on OGG1 protein expression suggest that it was physiologically regulated in response to oxidative modulation of ErbB, and that these might be dysregulated in the signaling pathway involving AKT in the progression of thyroid malignancies with RET/PTC rearrangements.

In the oil sector, a novelty in the centrifugal extraction system is represented by the multi-phase decanters (DMF) that work without adding process water and with the advantage of recovering a dried pomace and a by-product, called “pâté”, consisting of the pulp and its vegetation water, without traces of stone. The pâté has a high content of phenolic compounds, mainly represented by secoiridoids and verbascoside. The present work investigated the efficacy of two different ways of debittering (by sequential filtrations and spontaneous fermentation) of DMF pâté from three olive cultivars (Olea europaea L. “Leccino”, “Carboncella” and “Tortiglione”) to make the pâté edible, and, contemporary, investigated also the effect of its phenolic bioactive extracts on pathogenic bacteria and colon cancer cell model. Daily filtrations of pâté of the three cultivars have been shown to be more efficient in phenolic degradation. The activity of the indigenous microflora on the other hand takes a longer time to degrade the phenolic component and therefore to de-bitter it. None of pâté showed antibacterial activity. Colorimetric assay MTS for cell viability and metabolic activity tested on colon cancer cells Caco-2 and HCT116 suggest a potential beneficial effect of the dried extracts probably related to the modulation of gene expression under these treatments.

The identification of germline variants predisposing to hereditary nonpolyposis colorectal cancer (HNPCC) is crucial for clinical management of carriers, but several probands remain negative for such variants or bear variants of uncertain significance (VUS). Here we describe the results of integrative molecular analyses in 132 HNPCC patients providing evidences for improved genetic testing of HNPCC with traditional or next generation methods. Patients were screened for: germline allele-specific expression (ASE), nucleotide variants, rearrangements and promoter methylation of mismatch repair (MMR) genes; germline EPCAM rearrangements; tumor microsatellite instability (MSI) and immunohistochemical (IHC) MMR protein expression. Probands negative for pathogenic variants of MMR genes were screened for germline APC and MUTYH sequence variants. Most germline defects identified were sequence variants and rearrangements of MMR genes. Remarkably, altered germline ASE of MMR genes was detected in 8/22 (36.5%) probands analyzed, including 3 cases negative at other screenings. Moreover, ASE provided evidence for the pathogenic role and guided the characterization of a VUS shared by 2 additional probands. No germline MMR gene promoter methylation was observed and only one EPCAM rearrangement was detected. In several cases, tumor IHC and MSI diverged from germline screening results. Notably, APC or biallelic MUTYH germline defects were identified in 2/19 probands negative for pathogenic variants of MMR genes. Our results show that ASE complements gDNA-based analyses in the identification of MMR defects and in the characterization of VUS affecting gene expression, increasing the number of germline alterations detected. An appreciable fraction of probands negative for MMR gene variants harbors APC or MUTYH variants. These results indicate that germline ASE analysis and screening for APC and MUTYH defects should be included in HNPCC diagnostic algorithms.