A scanning electron microscopic study of the surface morphology of nuptial pads in male amphibians (Genus: Bombina, Pelophylax, Rana)

The fine structure of nuptial pad surface of the anuran amphibians Bombina variegata, Pelophylax epeiroticus, Pelophylax ridibundus and Rana dalmatina, was examined by scanning electron microscopy. Nuptial pads are cutaneous secondary sexual characters of males that appear during the breeding season and disappear afterwards following an annual cycle. In males of P. epeiroticus, P. ridibundus and R. dalmatina, nuptial pads were observed on the ventrolateral aspect of the first digit (the thumb) as darkish and remarkably keratinized papillae. In males of B. variegata nuptial pads were almost black and very visible on the thumb, the second and the third digit of the front legs. They also extended on the ventral surface of the forearms. Under scanning electron microscope numerous small papillae were observed rising above pad’s surface. In P. epeiroticus, P. ridibundus and R. dalmatina, these papillae were almost rounded at the base while at the dome shaped top they had many microprocesses organised in groups, thus assuming the shape of a “flower” which differed slightly among these three ranid species. In B. variegata the protuberances were conical with heavily keratinized hooks without microprocesses. Our results show that surface morphology of nuptial pads is unique for each species and could be considered as a species-specific character

Cytotopic (Cyto-) IL-15 as a New Immunotherapy for Prostate Cancer:Recombinant Production in Escherichia coli and Purification

Interleukin-15 (IL-15) is a cytokine previously suggested as a potential immunotherapy for cancer treatment. IL-15 can effectively reduce tumor growth in many preclinical tumor models including prostate cancer. This is due to its ability to expand and activate immune cells, such as CD8(+) T cells and natural killer cells. To increase the potency of IL-15, we have engineered a protein variant that can be modified to localize and be retained in tissues where it is administered. However, the production of recombinant IL-15, the purity, and correct refolding of the final protein is not always ideal. In the current study, we aimed to optimize the methodology for production and purification of a modified recombinant human IL-15 and investigate the efficacy of the produced protein in the treatment of prostate tumors. Human IL-15 with its polypeptide sequence modified at the C-terminus to enable thiol conjugation with membrane localizing peptides, was produced in E. coli and purified using mild denaturing conditions (2M urea) from a washing step or from solubilization of inclusion bodies. The purified protein from the wash fraction was conjugated to a myristoylated peptide to form a membrane-localizing IL-15 (cyto-IL-15). The efficacy of cyto-IL-15 was investigated in subcutaneous TRAMP-C2 prostate tumors in mice and compared with cyto-IL-15 derived from protein purified from inclusion bodies (cyto-IL-15 Gen). When mild denaturing conditions were used for purification, the largest amount of IL-15 was collected from the wash fraction and a smaller amount from inclusion bodies. The protein from the wash fraction was mainly present as a monomer, whereas the one from inclusion bodies formed homodimers and higher complexes. After cytotopic modification, the purified IL-showed great efficacy in delaying prostate tumor growth (∼50%) and increased mice survival by ∼1.8-fold compared with vehicle. This study demonstrates an alternative, inexpensive and efficient method to produce and purify a modified version of IL-15 using mild denaturing conditions. This IL-15, when cytotopically modified, showed great efficacy as a monotherapy in prostate tumors in mice further highlighting the potential of IL-15 as a cancer immunotherapy

Correlation of Ultrasound Shear Wave Elastography with Pathological Analysis in a Xenografic Tumour Model

The objective of this study was to evaluate the potential value of ultrasound (US) shear wave elastography (SWE) in assessing the relative change in elastic modulus in colorectal adenocarcinoma xenograft models in vivo and investigate any correlation with histological analysis. We sought to test whether non-invasive evaluation of tissue stiffness is indicative of pathological tumour changes and can be used to monitor therapeutic efficacy. US-SWE was performed in tumour xenografts in 15 NCr nude immunodeficient mice, which were treated with either the cytotoxic drug, Irinotecan, or saline as control. Ten tumours were imaged 48 hours post-treatment and five tumours were imaged for up to five times after treatment. All tumours were harvested for histological analysis and comparison with elasticity measurements. Elastic (Youngs) modulus prior to treatment was correlated with tumour volume (r = 0.37, p = 0.008). Irinotecan administration caused significant delay in the tumour growth (p = 0.02) when compared to control, but no significant difference in elastic modulus was detected. Histological analysis revealed a significant correlation between tumour necrosis and elastic modulus (r = -0.73, p = 0.026). SWE measurement provided complimentary information to other imaging modalities and could indicate potential changes in the mechanical properties of tumours, which in turn could be related to the stages of tumour development.Funding Agencies|Institute of Cancer Research; Engineering and Physical Sciences Research Council; Cancer Research UK Cancer Imaging Centre at the Institute of Cancer Research; NHS</p