Environmental assisted cracking of pipeline steels in CO2 containing environment

Buried pipelines are susceptible to Environmental Assisted Cracking (EAC) in three different conditions: Stress Corrosion Cracking in the presence of Carbonate-Bicarbonate (CB-SCC), Near Neutral Stress Corrosion Cracking (NN-SCC) and Hydrogen Embrittlement (HE). Both CB-SCC and NN-SCC involve in their mechanism the presence of CO2 and/or its dissociated species dissolved in moisture under the coating, while the generic term of HE refers to the phenomena of brittle fracture taking place for entry of atomic hydrogen inside the metal lattice, owing to the applied cathodic protection. Historically the CB-SCC was the first established form of stress corrosion on pipelines in the 60s, while Trans Canada Pipeline published in 1985-86 the first case of NN-SCC. The initiation conditions and the mechanism of propagation of CB-SCC were well established by means of the works of Parkins in 70ths. Many studies were carried out on NN-SCC, but actually there is not an exhaustive understanding of the problem. This paper summarizes the results obtained by authors on pipeline steels in NN-SCC promoting environments. Constant load, constant deformation, slow strain rate, slow bending and corrosion fatigue tests were executed on different grades of pipeline steels. The obtained results pointed out the effect of continuously plastic deformation to observe the propagation of NN-SCC cracks. NN-SCC cracks are preferentially nucleated from localized attacks. Inside the pits, the decreasing of pH enhances the hydrogen ions reduction. The results of slow strain rate tests are in agreement with a hydrogen embrittlement mechanism for the NN-SCC cracks propagation. Electrochemical tests, potentiodynamic and cyclic voltammetry were also carried out in order to analyze the effect of temperature, pH, CO2 and bicarbonate concentration on the pitting initiation. A pre-corrosion procedure, using a solution with high concentration of bicarbonate ions saturated with CO2 and a great number of voltammetry cycles was developed to obtain localized attacks on the specimen surface, similar than those observed in the failure analysis of the cracked pipelines. The SSR and slow bending tests carried out on these pre-corroded specimens evidenced presence of cracks with the same morphology of NN-SCC. Finally some corrosion-fatigue tests carried out on linear elastic fracture mechanics specimens (Single Edge Notch three four point Bending specimens) evidenced the increasing of fatigue propagation crack growth in NN-SCC environment. In the range of traditional and innovative pipeline steels, the mechanical properties (ultimate tensile strength and yield strength) seem do not influence their NN-SCC resistance

Environmentally assisted cracking of pipeline steels in CO2 containing environment at near-neutral pH

The paper summarizes the results obtained by authors concerning near-neutral stress corrosion cracking (SCC) phenomena on buried pipelines. Stress corrosion tests according to different methodologies and fatigue propagation tests were executed on traditional steel grades. Tests were performed also on specimens precorroded according to a procedure developed to produce localized attacks similar to those observed during failure analysis on pipelines, acting as preferential sites for crack nucleation. Electrochemical tests were carried out in order to evaluate the influence of environmental parameter on pit formation. The main role of continuous plastic deformations on near-neutral SCC is confirmed. The effect of pH, CO2, and bicarbonate concentrations on pitting formation is discussed

Geographical restriction of Hepatitis E virus circulation in wild boars (Sus scrofa) in Emilia-Romagna region, Northern Italy

Hepatitis E virus (HEV) is a single‐strand RNA virus that causes an acute viral hepatitis in humans. Among its eight recognized genotypes, HEV-3 and HEV-4 are zoonotic, infecting humans, pigs and feral pigs. Recently, HEV-3 has been also detected in red deer, which represents another reservoir of HEV. Consumption of raw pork products (mainly liver sausages), undercooked wild boar meat, raw wild boar liver and deer meat has been responsible for foodborne HEV human worldwide. From November 2018 to March 2019, liver samples collected from 97 wild boars hunted in Emilia-Romagna region (Northern Italy) were tested for HEV RNA. The hunting area included two territories for an extension of 33 km2, named A (about 13 km2, natural park, deciduous wood) and B (about 20 km2, cultivated fields in proximity of a river) areas. Distance between the two areas ranged between 8 to 10 km. A total of 73 wild boars were hunted in area A, and 24 in area B. HEV RNA was detected by Real‐time RT–PCR in 23/73 liver samples of wild boars living in area A only (31.5% - 95% CI: 22.0-42.8%). The HEV sequences (n=13) clustered within genotype 3. The majority of positives belonged to animals < 12 months (12/25; 48%), followed by subadults (13-24 months) (7/16; 43.8%) and adults (4/32; 12.5%). This difference was found to be statistically significant (p = 0.0024). In absence of pig farms, the restriction of HEV-positive animals to a well-defined territory of 13 km2 (Boschi di Carrega Regional Park) could hypothetically be related to the presence of red deer (Cervus elaphus), which lived in area A at the beginning of the hunting season. Further studies are needed to confirm or deny our hypothesis

PROTEASOME INHIBITORS MODULATE OSTEOCYTE DEATH AND AUTOPHAGY IN MULTIPLE MYELOMA.

Background: Cell death and autophagy are the main cellular processes involved in the regulation of bone remodeling by osteocytes. Recently we have demonstrated that an increased osteocyte death is involved in multiple myeloma (MM)-induced osteolysis through the upregulation of osteoclast recruitment. Aims: Because proteasome inhibitors including Bortezomib (BOR) are known to be able to target osteoblasts in this study we have investigated the potential effect of these drugs on osteocytes and their cell death and autophagy. Methods: Firstly the effect of the proteasome inhibitors BOR and MG262 on osteocyte viability was evaluated in vitro in murine osteocytic cell line MLO-Y4 and in the human pre-osteocytic one HOB-01. Both cell lines were co-coltured for 48 hours in the presence or absence of the human myeloma cell lines (HMCLs) RPMI8226 and JJN3, placed in a transwell insert in the presence or the absence of BOR or MG262. Moreover the effect of proteasome inhibitors on dexamethasone (DEX)-induced MLO-Y4 death, obtained at high doses (10-5-10-6M), was checked in combination with PTH(1-34). To evaluate the presence of autophagy and apoptosis in osteocytes, we checked the expression of both autophagic marker LC3 and apoptotic marker APAF-1 by confocal microscopy in the co-colture system with MLO-Y4 and RPMI-8226. Finally we performed a retrospective histological evaluation on bone biopsies of a cohort of 31 newly diagnosis MM underwent to different treatments including BOR-based regimen. Bone biopsies were obtained at the diagnosis and after an average time of 12 months of treatment. Osteocyte viability was evaluated in a total of 500 lacunae per histological sections. Results: The in vitro treatment with BOR or MG262 significantly blunted MLO-Y4 and HOB-01 cell death. Similarly, DEXinduced MLO-Y4 death was reduced by proteasome inhibitors. Interestingly, we found that both proteasome inhibitors potentiated the PTH (1-34) short-term effects on DEX-induced osteocyte death. Prevalence of autophagic cell death compared to apoptosis was observed in this system. In line with these data, we showed that neither the HMCLs nor treatment with DEX increase the apoptotic death and caspase 3 activation in both MLO-Y4 and HOB-01 cell lines. BOR treatment increased the basal level of LC3 indicating a pro-survival and protective function of autophagy against the BOR-induce stress. On the contrary, when the cells undergo to a stronger stress such as in the presence of HMCLs or by treatment with high dose of DEX we found that both proteasome inhibitors blocked autophagic cell death in osteocytes. In the in vivo study we found a significant increase of the number of viable osteocytes in MM patients treated with BOR-based regimen as compared to those treated without BOR (% median increase: +6% vs. +1.30%; p=0.017). Patients treated with BOR alone showed the highest increase of osteocyte viability, as compared to those either treated without BOR (+11.6% vs. +1.3%, p=0.0019) or treated with BOR plus DEX (+11.6% vs. +4.4%, p=0.01). On the other hand, any significant difference was not observed in patients treated with Thalidomide (THAL) or Immunomodulatory drugs (IMiDs) than in those untreated with these drugs (p= 0.7). A multiple regression non-parametric analysis showed that BOR had a significant positive impact on osteocyte viability (p=0.042) whereas THAL/IMiDs as well as Zoledronic acid (ZOL) treatments have not (p=0.2). BOR also counterbalanced the negative effect of DEX treatment (p=0.035). Summary/Conclusion: Our data suggest that proteasome inhibitors blunted osteocyte cell death induced by MM cells and DEX through the modulation of the autophagy and potentiated the effect of PTH. Overall our in vitro and in vivo data support the use of BOR to improve bone integrity in MM patients

Mechanism of Action of Bortezomib and the New Proteasome Inhibitors on Myeloma Cells and the Bone Microenvironment: Impact on Myeloma-Induced Alterations of Bone Remodeling

Multiple myeloma (MM) is characterized by a high capacity to induce alterations in the bone remodeling process. The increase in osteoclastogenesis and the suppression of osteoblast formation are both involved in the pathophysiology of the bone lesions in MM. The proteasome inhibitor (PI) bortezomib is the first drug designed and approved for the treatment of MM patients by targeting the proteasome. However, recently novel PIs have been developed to overcome bortezomib resistance. Interestingly, several preclinical data indicate that the proteasome complex is involved in both osteoclast and osteoblast formation. It is also evident that bortezomib either inhibits osteoclast differentiation induced by the receptor activator of nuclear factor kappa B (NF-κB) ligand (RANKL) or stimulates the osteoblast differentiation. Similarly, the new PIs including carfilzomib and ixazomib can inhibit bone resorption and stimulate the osteoblast differentiation. In a clinical setting, PIs restore the abnormal bone remodeling by normalizing the levels of bone turnover markers. In addition, a bone anabolic effect was described in responding MM patients treated with PIs, as demonstrated by the increase in the osteoblast number. This review summarizes the preclinical and clinical evidence on the effects of bortezomib and other new PIs on myeloma bone disease

Klebsiella pneumoniae carrying multiple alleles of antigen 43-encoding gene of Escherichia coli associated with biofilm formation

A clinical strain of Klebsiella pneumoniae typed as sequence type 307 carrying three different alleles of the flu gene encoding the Escherichia coli virulence factor antigen 43 associated with biofilm formation was detected and characterized. The flu alleles are located in the chromosome inside putative integrative conjugative elements. The strain displays the phenotypes associated with Ag43, i.e. bi-phasic colony morphology and enhanced biofilm production. Furthermore, the strain produces low amount of capsule known to affect Ag43 function. Analysis of 1431 worldwide deposited genomes revealed that 3.7% Klebsiella pneumoniae carry one or two flu alleles

New Insights into Osteogenic and Chondrogenic Differentiation of Human Bone Marrow Mesenchymal Stem Cells and Their Potential Clinical Applications for Bone Regeneration in Pediatric Orthopaedics

Human mesenchymal stem cells (hMSCs) are pluripotent adult stem cells capable of being differentiated into osteoblasts, adipocytes, and chondrocytes. The osteogenic differentiation of hMSCs is regulated either by systemic hormones or by local growth factors able to induce specific intracellular signal pathways that modify the expression and activity of several transcription factors. Runt-related transcription factor 2 (Runx2) and Wnt signaling-related molecules are the major factors critically involved in the osteogenic differentiation process by hMSCs, and SRY-related high-mobility-group (HMG) box transcription factor 9 (SOX9) is involved in the chondrogenic one. hMSCs have generated a great interest in the field of regenerative medicine, particularly in bone regeneration. In this paper, we focused our attention on the molecular mechanisms involved in osteogenic and chondrogenic differentiation of hMSC, and the potential clinical use of hMSCs in osteoarticular pediatric disease characterized by fracture nonunion and pseudarthrosis

New Insights into Osteogenic and Chondrogenic Differentiation of Human Bone Marrow Mesenchymal Stem Cells and Their Potential Clinical Applications for Bone Regeneration in Pediatric Orthopaedics

Human mesenchymal stem cells (hMSCs) are pluripotent adult stem cells capable of being differentiated into osteoblasts, adipocytes, and chondrocytes. The osteogenic differentiation of hMSCs is regulated either by systemic hormones or by local growth factors able to induce specific intracellular signal pathways that modify the expression and activity of several transcription factors. Runt-related transcription factor 2 (Runx2) and Wnt signaling-related molecules are the major factors critically involved in the osteogenic differentiation process by hMSCs, and SRY-related high-mobility-group (HMG) box transcription factor 9 (SOX9) is involved in the chondrogenic one. hMSCs have generated a great interest in the field of regenerative medicine, particularly in bone regeneration. In this paper, we focused our attention on the molecular mechanisms involved in osteogenic and chondrogenic differentiation of hMSC, and the potential clinical use of hMSCs in osteoarticular pediatric disease characterized by fracture nonunion and pseudarthrosis