Human chondrocytes and mesenchymal stem cells grown onto engineered scaffold

The development of improved methods for treatment of chondral defects using autologous cells in combination with biomaterials leads to a new generation of implantable devices. Their association gives rise to a hybrid construct combining biological and material components that can be specifically committed. The comprehension of cellular and molecular mechanisms of cartilage repair and the use of biomaterials in combination with chondrocytes or mesenchymal stem cells in the treatment of cartilage defects has opened a new era of therapeutical strategies. Recently, their applicability in the treatment of early lesions in osteoarthritis is under investigation. To obtain new information on the behaviour of chondrocytes and mesenchymal stem cells grown on a hyaluronan derivative scaffold (Hyaff-11) already used in cartilage repair, we analysed a series of molecules expressed by these cells by Real-Time RT-PCR and immunohistochemical analyses. The data obtained with this work showed that this biomaterial is able to reduce the expression of some catabolic molecules by human chondrocytes and provide a good environment to support the differentiation of mesenchymal stem cells in chondrogenic sense. These observations confirm Hyaff-11 as a suitable scaffold both for chondrocytes and mesenchymal stem cells for the treatment of articular cartilage defects.     

Diffuse cutaneous candidiasis in a dog. Diagnosis by PCR-REA

The authors describe a clinical case of cutaneous candidiasis in a dog with dermatological lesions, characterized by persistent alopecia, crusts, ulcers and scales. Predisposing factors such as the use of corticosteroids, the concomitan presence of an autoimmune disease (pemphigus foliaceus) and an infection of ehrlichiosis caused by Ehrlichia canis were observed. Histopathological findings included signs of orthokeratotic hyperkeratosis, moderate follicular keratosis and light epidermic acanthosis. The reactive process included an infiltrative superficial dermatitis and a mural folliculitis with prevalent participation of macrophages and lymphocytes. The application of PCR-Restriction Enzyme Analysis (REA) method on cutaneous specimens in veterinary medicine is an extremely interesting diagnostic tool. Its use, together with other techniques, such as mycologic, cytologic and histological examinations, allowed us to identify Candida albicans as aetiological agent in this particular case.     

Oncosuppressor methylation: a possible key role in colon metastatic progression

K‐RAS and BRAF gene mutations are mandatory to set anti‐EGFR therapy in metastatic colorectal cancer (mCRC) patients. Due to the relationship of these mutations with tumor epigenotype, we hypothesized the potential role of oncosuppressor methylation of genes involved in K‐RAS/BRAF pathway (CDKN2A, RASSF1A, and RARbeta suppressor genes) in inhibiting EGFR signaling cascade. Primary tumor and synchronous liver metastatic tissues of 75 mCRC patients were characterized for promoter methylation by QMSP and for K‐RAS and BRAF mutations. RARbeta, RASSF1A, and CDKN2A genes were methylated in 82%, 35%, and 26% of primary tumors, respectively. RASSF1A resulted significantly more frequently methylated in liver metastasis than in primary site (P = 0.015), while RARbeta was significantly lower methylated in distant metastasis (P = 1.2 × 10^?6). As regards methylation content, RASSF1A methylation status was significantly higher in liver metastasis with respect to primary tumor (P = 0.000) underlying the role of this gene in liver metastatic progression. In our series K‐RAS and BRAF were mutated in 39% and 4% of cases, respectively. Methylation frequencies seemed to be unrelated to gene mutations; on the other hand, RASSF1A mean content methylation resulted significantly higher in liver than in primary tumor (288.78 vs. 56.23, respectively, P = 0.05) only in K‐RAS wild‐type cases sustaining a specific role of this gene in metastatic site thus supporting its function in strengthening the apoptotic role of K‐RAS. These evidences held the role of oncosuppressor methylation in both colon tumorigenesis and progression and suggested that epigenetic events should be taken into account when biological therapies in mCRC patients have to be set. J. Cell. Physiol. 226: 1934–1939, 2011. © 2010 Wiley‐Liss, Inc.     

Slime-producing Staphylococcus epidermidis and S. aureus in acute bacterial conjunctivitis in soft contact lens wearers

In recent years, an increase in ocular pathologies related to soft contact lens has been observed. The most common infectious agents were Staphylococcus spp. Some strains produce an extracellular polysaccharidic slime that can cause severe infections. Polysaccharide synthesis is under genetic control and involves a specific intercellular adhesion (ica) locus, in particular, icaA and icaD genes. Conjunctival swabs from 97 patients with presumably bacterial bilateral conjunctivitis, wearers of soft contact lenses were examined. We determined the ability of staphylococci to produce slime, relating it to the presence of icaA and icaD genes. We also investigated the antibiotic susceptibility and Pulsed Field Gel Electrophoresis (PFGE) patterns of the clinical isolates. We found that 74.1% of the S. epidermidis strains and 61.1% of the S. aureus strains isolated were slime producers and showed icaA and icaD genes. Both S. epidermidis and S. aureus slime-producing strains exhibited more surface hydrophobicity than non-producing slime strains. The PFGE patterns overlapped in S. epidermidis strains with high hydrophobicity. The similar PFGE patterns were not related to biofilm production. We found scarce matching among the Staphylococcus spp. studied, slime production, surface hydrophobicity and antibiotic susceptibility.     

Discovery of a regular nesting area of loggerhead turtle <Emphasis Type="Italic">Caretta caretta</Emphasis> in southern Italy: a new perspective for national conservation

Loggerhead turtle Caretta caretta nesting in Italy had been reported to be limited to the Pelagian Islands and only sporadically elsewhere. As presence of loggerhead turtle nests had occasionally been reported (1988–1999) along about 200 km of the Ionian coast of Calabria, we carried out a project to assess the actual state of the nesting population between 2000 and 2004. We divided the coastline in two sectors (A: 52 km, and B: 146 km) that were monitored from mid-June to end of July for a total of n = 174 monitoring days and 1,813.6 km patrolled on foot with different intensities (extensive versus intensive). In sector B, through extensive monitoring we did not find any emergence tracks, but in sector A by intensive survey (2002–2004: one survey/3.64 days) we detected 3–8 nests/year. In total, 25 nests (both observed and reported), were recorded in our study area, and an assessment of a total of 15–16 nests/year was suggested. These figures, within the national scenario depicted from the review of known nesting events in the last 40 years (88 records concerning more than 143–144 nests), show that loggerhead turtle nesting has been underestimated in Italy, due to inadequate monitoring protocols, and that nesting is more frequent than expected (at least 30–40 nests/year). Conservation strategies in Italy should then focus not only on the reduction of mortality at sea, but also include large-scale actions to preserve scattered (but regular) nesting events.     

High‐resolution crystal structure of human Mapkap kinase 3 in complex with a high affinity ligand

The Mapkap kinases 2 and 3 (MK2 and MK3) have been implicated in intracellular signaling pathways leading to the production of the pro‐inflammatory cytokine tumor necrosis factor alpha. MK2 has been pursued by the biopharmaceutical industry for many years for the development of a small molecule anti‐inflammatory treatment and drug‐like inhibitors have been described. The development of some of these compounds, however, has been slowed by the absence of a high‐resolution crystal structure of MK2. Herein we present a high‐resolution (1.9 Å) crystal structure of the highly homologous MK3 in complex with a pharmaceutical lead compound. While all of the canonical features of Ser/Thr kinases in general and MK2 in particular are recapitulated in MK3, the detailed analysis of the binding interaction of the drug‐like ligand within the adenine binding pocket allows relevant conclusions to be drawn for the further design of potent and selective drug candidates.     

Biomarkers in malignant mesothelioma: diagnostic and prognostic role of soluble mesothelin-related peptide

Soluble mesothelin-related peptide (SMRP) is a biomarker that has been proposed for differential diagnosis from pleural metastatic cancer, as well as prognosis and treatment monitoring of malignant pleural mesothelioma (MM). The aim of this study was to evaluate the role of SMRP in clinic management of MM. We assayed the SMRP concentrations in 354 subjects: 109 healthy volunteers with no history of exposure to asbestos, 26 patients with previous occupational asbestos exposure but who were free from pleural or parenchymal disease, 48 patients with asbestosis, 110 patients with pleural plaques, 25 patients with lung cancer, and 36 patients with MM. We also tested SMRP titers in 2 patients with MM at 5 different times of the disease, to evaluate the trend of the biomarker in the course of therapy. Our data confirm previous experiences with the use of SMRP as a diagnostic marker of MM. Low SMRP levels at diagnosis seem to have a positive prognostic significance.     

Different methylation and MicroRNA expression pattern in male and female familial breast cancer

Epigenetic regulation, has been very scarcely explored in familial breast cancer (BC). In the present study RASSF1A and RAR beta promoter methylation and miR17, miR21, miR 124, and let‐7a expression were investigated to highlight possible differences of epigenetic regulation between male and female familial BC, also in comparison with sporadic BC. These epigenetic alterations were studied in 56 familial BC patients (27 males and 29 females) and in 16 female sporadic cases. RASSF1A resulted more frequently methylated in men than women (76% vs. 28%, respectively, P = 0.0001), while miR17 and let‐7a expression frequency was higher in women than in men (miR17: 66% in women vs. 41% in men, P < 0.05; let‐7a: 45% in women vs. 15% in men, P = 0.015). RASSF1A methylation affected 27.6% of familial BC while 83% of familial cases showed high expression of the gene (P = 0.025); on the contrary, only 17% of familial BC presented RAR beta methylation and 55% of familial cases overexpressed this gene (P = 0.005). Moreover, miR17, miR21, and let‐7a resulted significantly overexpressed in familial compared to sporadic BC. RASSF1A overexpression (86% vs. 65%, P = 0.13) and RAR beta overexpression (57% vs. 32%, P = 0.11) were higher in BRCA1/2 carriers even if not statistical significance was reached. BRCA mutation carriers also demonstrated significant overexpression of: miR17 (93% vs. 35%, P = 0.0001), let‐7a (64% vs. 16%, P = 0.002), and of miR21 (100% vs. 65%, P = 0.008). In conclusion, the present data suggest the involvement of RASSF1A in familial male BC, while miR17 and let‐7a seem to be implied in familial female BC. J. Cell. Physiol. 228: 1264–1269, 2013. © 2012 Wiley Periodicals, Inc.     

Changes in vascular and transpiration flows affect the seasonal and daily growth of kiwifruit (Actinidia deliciosa) berry

Background and Aims

The kiwifruit berry is characterized by an early stage of rapid growth, followed by a relatively long stage of slow increase in size. Vascular and transpiration flows are the main processes through which water and carbon enter/exit the fruit, determining the daily and seasonal changes in fruit size. This work investigates the biophysical mechanisms underpinning the change in fruit growth rate during the season.

Methods

The daily patterns of phloem, xylem and transpiration in/outflows have been determined at several stages of kiwifruit development, during two seasons. The different flows were quantified by comparing the diurnal patterns of diameter change of fruit, which were then girdled and subsequently detached while measurements continued. The diurnal courses of leaf and stem water potential and of fruit pressure potential were also monitored at different times during the season.

Key Results

Xylem and transpiration flows were high during the first period of rapid volume growth and sharply decreased with fruit development. Specific phloem import was lower and gradually decreased during the season, whereas it remained constant at whole-fruit level, in accordance with fruit dry matter gain. On a daily basis, transpiration always responded to vapour pressure deficit and contributed to the daily reduction of fruit hydrostatic pressure. Xylem flow was positively related to stem-to-fruit pressure potential gradient during the first but not the last part of the season, when xylem conductivity appeared to be reduced.

Conclusions

The fruit growth model adopted by this species changes during the season due to anatomical modifications in the fruit features.