Human fibroblasts from normal and malignant breast tissue grown in vitro show a distinct senescence profile and telomerase activity

The telomerase activity and the senescence profile of cultured breast fibroblasts from normal human interstitial and malignant stromal tissue were studied in comparison with their proliferation and differentiation pattern. Fibroblasts were grown either in the presence or absence of a conditioned medium (CM) obtained from cultures of the oestrogen receptor-positive breast cancer MCF-7 cell line. At different passages (from the 2nd up to the 48th), fibroblasts were examined for the telomerase activity by the Telomerase Repeats Amplification Protocol (TRAP) assay, for proliferation profile by Ki-67 antigen expression, and the myofibroblast or smooth muscle cell-like differentiation pattern by immunofluorescence with monoclonal antibodies specific for smooth muscle markers. Serial passages of fibroblasts from normal or tumour breast reveal that the relationship between the levels of telomerase activity and phenotypic/proliferation profile changes with cell subcultivation in a different manner in the two cell populations. The fibroblasts from normal tissue completed 12 passages in a CM-independent way prior to senescence whereas fibroblasts from tumour stroma senescence were attained after 48 passages. These cells showed a marked decrease of telomerase activity, growth rate and smooth muscle -actin expressing myofibroblasts after the 32nd passage. CM treatment of this fibroblast population induces a decline in the myofibroblast content, which precedes the changes in telomerase activity. Passaged fibroblasts from normal breast tissue can be converted to myofibroblasts upon CM treatment whereas those from tumour stroma were CM-insensitive. Taken together our data suggest that a heterogeneous fibroblast population with different life span is activated/recruited in the breast interstitium and poses the problem of a unique activation/recruitment of fibroblasts in neoplastic conditions.     

Antifouling activity of enzyme‐functionalized silica nanobeads

The amelioration of biofouling in industrial processing equipment is critical for performance and reliability. While conventional biocides are effective in biofouling control, they are potentially hazardous to the environment and in some cases corrosive to materials. Enzymatic approaches have been shown to be effective and can overcome the disadvantages of traditional biocides, however they are typically uneconomic for routine biofouling control. The aim of this study was to design a robust and reusable enzyme‐functionalized nano‐bead system having biofilm dispersion properties. This work describes the biochemical covalent functionalization of silica‐based nanobeads (hereafter referred to as Si‐NanoB) with Proteinase K (PK). Results showed that PK‐functionalized Si‐NanoB are effective in dispersing both protein‐based model biofilms and structurally altering Pseudomonas fluorescens biofilms, with significant decreases in surface coverage and thickness of 30.1% and 38.85%, respectively, while increasing surface roughness by 19 % following 24 h treatments on bacterial biofilms. This study shows that enzyme‐functionalized nanobeads may potentially be an environmentally friendly and cost effective alternative to pure enzyme and chemical treatments. Biotechnol. Bioeng. 2016;113: 501–512. © 2015 The Authors. Biotechnology and Bioengineering Published by Wiley Periodicals, Inc.     

Age-dependent changes in the mechanical properties of tail tendons in TGF-beta inducible early gene-1 knockout mice.

The purpose of this study is to investigate age-dependent changes in the architecture and mechanical properties of tendon in TGF-beta inducible early gene-1 (TIEG) knockout mice. Wild-type and TIEG knockout mice, aged 1, 2, and 15 mo, were used. The mechanical properties of tail tendons isolated from these mice were determined using uniaxial tensile ramp (0.05 mm/s) and relaxation (5 mm/s) tests, with a strain of 10%. Mechanical parameters (Young's modulus from the ramp test; fast and static stresses from the relaxation test) were measured and recorded. The structure of the tail tendon fascicle was characterized by transmission electron microscopy. The results of the mechanical testing revealed no significant difference between the knockout and wild-type groups at 1 or 15 mo of age. However, the fascicles of the knockout mice at 3 mo of age exhibited decreased fast and static stresses compared with those of the wild-type mice. Electron microscopy revealed an increase in fibril size in the knockout mouse tendons relative to wild-type controls at 1 and 3 mo of age. These data indicate an important role for TIEG in tendon microarchitecture and strength in adult mice.     

Angiosuppression and chemotherapy: strategies aimed at their integration in cancer patients

A number of antiangiogenic agents have been developed as pharmaceuticals and are currently being tested in clinical studies. Potential strategies to enhance the activity of angiogenesis inhibitors could be to combine them, or better still, to administer them either sequentially or concurrently with cytotoxic drugs. Chemotherapy would be a more appropriate initial choice for patients with advanced disease since cytostatic agents can induce a fast regression of the tumor and cancer-related symptoms. Antiangiogenic treatment could be used after chemotherapy in patients who achieve disease remission to prolong the time to progression, the symptom-free interval and the overall survival. Antiangiogenic treatment is likely to attain an important role in the adjuvant setting. In fact, it could be used for prolonged periods after radical surgery to maintain dormancy of residual tumor cells. In spite of these promising preclinical data, several points need to be clarified before the initiation of clinical trials. In fact, certain misconceptions may interfere with their optimum design and result analysis.     

Electrospun micro- and nanofiber tubes for functional nervous regeneration in sciatic nerve transections

Background

Rhipicephalus (Boophilus) spp. ticks economically impact on cattle production in Africa and other tropical and subtropical regions of the world. Tick vaccines constitute a cost-effective and environmentally friendly alternative to tick control. The R. microplus Bm86 protective antigen has been produced by recombinant DNA technology and shown to protect cattle against tick infestations.

Results

In this study, the genes for Bm86 (R. microplus), Ba86 (R. annulatus) and Bd86 (R. decoloratus) were cloned and characterized from African or Asian tick strains and the recombinant proteins were secreted and purified from P. pastoris. The secretion of recombinant Bm86 ortholog proteins in P. pastoris allowed for a simple purification process rendering a final product with high recovery (35–42%) and purity (80–85%) and likely to result in a more reproducible conformation closely resembling the native protein. Rabbit immunization experiments with recombinant proteins showed immune cross-reactivity between Bm86 ortholog proteins.

Conclusion

These experiments support the development and testing of vaccines containing recombinant Bm86, Ba86 and Bd86 secreted in P. pastoris for the control of tick infestations in Africa.     

Filamentous fungal biofilm for production of human drug metabolites

In drug development, access to drug metabolites is essential for assessment of toxicity and pharmacokinetic studies. Metabolites are usually acquired via chemical synthesis, although biological production is potentially more efficient with fewer waste management issues. A significant problem with the biological approach is the effective half-life of the biocatalyst, which can be resolved by immobilisation. The fungus Cunninghamella elegans is well established as a model of mammalian metabolism, although it has not yet been used to produce metabolites on a large scale. Here, we describe immobilisation of C. elegans as a biofilm, which can transform drugs to important human metabolites. The biofilm was cultivated on hydrophilic microtiter plates and in shake flasks containing a steel spring in contact with the glass. Fluorescence and confocal scanning laser microscopy revealed that the biofilm was composed of a dense network of hyphae, and biochemical analysis demonstrated that the matrix was predominantly polysaccharide. The medium composition was crucial for both biofilm formation and biotransformation of flurbiprofen. In shake flasks, the biofilm transformed 86 % of the flurbiprofen added to hydroxylated metabolites within 24 h, which was slightly more than planktonic cultures (76 %). The biofilm had a longer effective lifetime than the planktonic cells, which underwent lysis after 2?×?72 h cycles, and diluting the Sabouraud dextrose broth enabled the thickness of the biofilm to be controlled while retaining transformation efficiency. Thus, C. elegans biofilm has the potential to be applied as a robust biocatalyst for the production of human drug metabolites required for drug development.     

Length-weight relationship of fish species from Central Amazon floodplain

Length-weight relationship (LWR) for 39 freshwater fish species captured in the Catalão Lake, a floodplain area at the confluence of the Amazonas and Negro rivers is presented. LWRs were calculated based on fish sampled over 18 years (1999–2017) using a set of ten gill nets with different mesh sizes, monthly immersed in water for 24-hr with 6-hr interval catches. Measurements were done for standard length (SL – 0.1 cm precision) and total weight (TW – 0.01 g precision). The LWRs were calculated by the linear regression of natural log-transformed SL and TW data: TW = a × SL^b. All statistical analyses were performed with R software. From all species considered, 32 are new LWR records for the international literature as well as for the Amazonian ichythyofauna; additional records expand the known size range for seven species.     

Endoscopic Endonasal Approach in the Management of Rathke’s Cleft Cysts

Objective

Rathke’s cleft cysts (RCCs) are quite uncommon sellar lesions that can extend or even arise in the suprasellar area. The purpose of this study is to evaluate the effectiveness of both standard and extended endoscopic endonasal approaches in the management of different located RCCs.

Methods

We retrospectively analyzed a series of 29 patients (9 males, 20 females) complaining of a RCC, who underwent a standard or an extended endoscopic transsphenoidal approach at the Division of Neurosurgery, Department of Neurosciences and Reproductive and Odontostomatological Sciences, of the Università degli Studi di Napoli "Federico II”. Data regarding patients’ demographics, clinical evaluation, cyst characteristics, surgical treatments, complications and outcomes were extracted from our electronic database (Filemaker Pro 11, File Maker Inc., Santa Clara, California, USA).

Results

A standard transsphenoidal approach was used in 19 cases, while the extended variation of the approach in 10 cases (5 purely suprasellar and 5 intra-suprasellar RCC). Cysts contents was fully drained in all the 29 cases, whilst a gross total removal, that accounts on the complete cyst wall removal, was achieved in an overall 55,1% of patients (16/29), specifically 36,8% (7/19) that received standard approach and 90% (9/10) of those that underwent to extended approach. We reported a 56.2% of recovery from headache, 38.5% of complete recovery and 53.8% of improvement from visual field defect and an overall 46.7% of improvement of the endocrine functions. Postoperative permanent DI rate was 10.3%, overall post-operative CSF leak rate 6.9%; recurrence/regrowth occurred in 4 patients (4/29, 13.8%), but only one required a second surgery.

Conclusion

The endoscopic transsphenoidal approach for the removal of a symptomatic RCC offers several advantages in terms of visualization of the surgical field during both the exposure and removal of the lesion. The “extended” variation of the endoscopic approach provides a direct access to the supradiaphragmatic space, allowing adequate view and room for the safe removal of selected supradiaphragmatic RCCs, regardless of the sellar size (even a not enlarged sella), and provides a higher likelihood of preserving normal pituitary tissue and functions.