Luca Ventura

Proline/arginine-rich end leucine-rich repeat protein N-terminus is a novel osteoclast antagonist that counteracts bone loss

hbdPRELP is a peptide corresponding to the N‐terminal heparin binding domain of the matrix protein proline/arginine‐rich end leucine‐rich repeat protein (PRELP). hbdPRELP inhibits osteoclastogenesis entering pre‐fusion osteoclasts through a chondroitin sulfate– and annexin 2–dependent mechanism and reducing the nuclear factor‐κB transcription factor activity. In this work, we hypothesized that hbdPRELP could have a pharmacological relevance, counteracting bone loss in a variety of in vivo models of bone diseases induced by exacerbated osteoclast activity. In healthy mice, we demonstrated that the peptide targeted the bone and increased trabecular bone mass over basal level. In mice treated with retinoic acid to induce an acute increase of osteoclast formation, the peptide consistently antagonized osteoclastogenesis and prevented the increase of the serum levels of the osteoclast‐specific marker tartrate‐resistant acid phosphatase. In ovariectomized mice, in which osteoclast activity was chronically enhanced by estrogen deficiency, hbdPRELP counteracted exacerbated osteoclast activity and bone loss. In mice carrying osteolytic bone metastases, in which osteoclastogenesis and bone resorption were enhanced by tumor cell–derived factors, hbdPRELP reduced the incidence of osteolytic lesions, both preventively and curatively, with mechanisms involving impaired tumor cell homing to bone and tumor growth in the bone microenvironment. Interestingly, in tumor‐bearing mice, hbdPRELP also inhibited breast tumor growth in orthotopic sites and development of metastatic disease in visceral organs, reducing cachexia and improving survival especially when administered preventively. hbdPRELP was retained in the tumor tissue and appeared to affect tumor growth by interacting with the microenvironment rather than by directly affecting the tumor cells. Because safety studies and high‐dose treatments revealed no adverse effects, hbdPRELP could be employed as a novel biological agent to combat experimentally induced bone loss and breast cancer metastases, with a potential translational impact.

HLA-DRB genotyping of an Italian mummy from the 16th century with signs of rheumatoid arthritis

Some paleopathological studies suggest that rheumatoid arthritis originated in the New World (among ancient Native Americans in Tennessee and neighbouring areas, 5000–500 BC); only after the discovery of America was the origin of the disease attributed to the Old World.1 In 1996, under the floors of the San Francisco church (Arezzo, Italy), a female mummy was discovered. This 50–55-year-old woman, re-named the “Braids Lady”, died at the end of the 16th century. Several diagnostic examinations disclosed distinctive rheumatoid arthritis skeletal deformities due to rheumatoid arthritis in her body: large erosions of the metacarpophalangeal joints of the left hand; lateral deviation of all the fingers, with a typical “Z” deformation of the first …

Effective Small Interfering RNA Therapy to Treat CLCN7-dependent Autosomal Dominant Osteopetrosis Type 2

In about 70% of patients affected by autosomal dominant osteopetrosis type 2 (ADO2), osteoclast activity is reduced by heterozygous mutations of the CLCN7 gene, encoding the ClC-7 chloride/hydrogen antiporter. CLCN7G215R-, CLCN7R767W-, and CLCN7R286W-specific siRNAs silenced transfected mutant mRNA/EGFP in HEK293 cells, in RAW264.7 cells and in human osteoclasts, with no change of CLCN7WT mRNA and no effect of scrambled siRNA on the mutant transcripts. Osteoclasts from Clcn7G213R ADO2 mice showed reduced bone resorption, a condition rescued by Clcn7G213R-specific siRNA. Treatment of ADO2 mice with Clcn7G213R-specific siRNA induced increase of bone resorption variables and decrease of trabecular bone mass, leading to an overall improvement of the osteopetrotic bone phenotype. Treatment did not induce overt adverse effects and was effective also with siRNAs specific for other mutants. These results demonstrate that a siRNA-based experimental treatment of ADO2 is feasible, and underscore a translational impact for future strategy to cure this therapeutically neglected form of osteopetrosis.

Genetic predisposition to rheumatoid arthritis in a Tuscan (Italy) ancient human remain.

Rheumatoid arthritis (RA) is currently believed to have originated in America, and after the discovery of this continent in 1492, to have been exported to the Old World. We evaluated the genetic predisposition to RA in the “Braids Lady” from Arezzo (Italy), a partially mummified womans body dating back to the end of 1500 AD which presents the anatomical and pathological features of this disease. The study of the polymorphic HLA-DRB1 locus, which includes alleles strongly associated with RA onset, has received much attention over recent years, especially the loci codifying for the DR1 and DR4 antigens, widely represented in the Mediterranean population, and for DR14, widespread among Native Americans. Molecular analysis was performed on extracts of DNA from the mummy, firstly from histological bone sections and then from the whole bone. Two different HLA typing techniques, PCR-sequence-specific oligonucleotides (PCR-SSO) and PCR-sequence-specific primers (PCR-SSP), were employed to identify HLA-DRB alleles. Both genotyping methods showed that the “Braids Lady” carried the DRB1*0101 allele, the serological equivalent of the DR1 antigen. Although the possession of RA risk factor genes cannot be considered a diagnostic marker, the positive result of the Italian mummy for DRB1*0101 and the RA features present, support the idea that this pathology was present in the Old World from at least the mid-16th century. A pathogenetic hypothesis of RA which might well explain its worldwide diffusion is the “molecular mimicry”, resulting from a cross-reactive antibody response between certain microbial antigens and shared epitopes of specific HLA-DR1, DR4 and DR14 susceptibility alleles, the frequency of which varies among different ethnic groups.

The Cutaneous Cancer of Ferdinando Orsini, 5th Duke of Gravina

18. Hauschild A, Popp G, Stockfleth E, et al. Effective photodynamic therapy of actinic keratoses on the head and face with a novel, self-adhesive 5-aminolaevulinic acid patch. Exp Dermatol. 2009;18(2):116-121. 19. Mamalis A, Koo E, Sckisel GD, Siegel DM, Jagdeo J. Temperature-dependent impact of thermal aminolaevulinic acid photodynamic therapy on apoptosis and reactive oxygen species generation in human dermal fibroblasts. Br J Dermatol. 2016;175(3):512-519.

Renal Calculosis of Pandolfo III Malatesta (1370-1427)

Renal Calculosis of Pandolfo III Malatesta (1370-1427) Valentina Giuffra, PhD, Luca Ventura, MD, Simona Minozzi, PhD, Agata Lunardini, Raimondo Quaresima, Lorenzo Arrizza, Gino Fornaciari, MD Division of Paleopathology, History of Medicine and Bioethics, Department of Oncology, Transplants and Advanced Technologies in edicine, University of Pisa, Italy; Department of Pathology, San Salvatore Hospital, L’Aquila, Italy; Department of Chemistry, hemical Engineering and Materials, University of L’Aquila, Italy; Centre of Electronic Microscopy, University of L’Aquila, Italy.

Hair granuloma of the prostate. A clinically silent, under-recognized complication of needle core biopsy

b-catenin in high-grade neuroendocrine tumours. These included large cell neuroendocrine tumours and small cell carcinoma. In carcinoid tumours, E-cadherin immunostaining was present in seven out of 16 TCs and five out of eight ACs. An impaired E-cadherin expression pattern was strongly correlated with the presence of nodal metastasis in all neuroendocrine tumours, although the difference in the carcinoid group did not reach statistical significance. This study was done on frozen material. The second study showed that a disarrayed E-cadherin or b-catenin pattern was strongly associated with lymph node metastasis in patients with AC. Our findings strongly support the theory that inhibition of the epithelium specific cell–cell adhesion molecule, E-cadherin, plays a significant role in the ability of lung carcinoid tumours to progress and, more importantly, to metastasize. With the help of markers we have tried to show that there are marked biological differences between TC and AC and there are subtle differences between TC with and without metastasis. With inclusion of more markers and a larger number of cases, the impact of this study might be enhanced.

Expression of Peroxisome Proliferator-Activated Receptor Alpha (PPARα) in Non-Somatotroph Pituitary Tumours and the Effects of PPARα Agonists on MMQ Cells

Peroxisome proliferator-activated receptor alpha (PPARα) has been involved in the regulation of somatotroph tumour cells and may be targeted by different drugs, some of them are in current clinical use. The aim of this study was to investigate the expression of PPARα in additional phenotypes of pituitary adenomas (PA), the relationship between PPARα and its potential molecular partner aryl hydrocarbon receptor interacting protein (AIP) in these tumours, and the effects of PPARα agonists on lactotroph cells. Seventy-five human PA - 57 non-functioning (NFPA) and 18 prolactinomas (PRL-PA) - were characterised for PPARα and AIP expression by real time RT-PCR and/or immunohistochemistry (IHC), and the effects of fenofibrate and WY 14 643 on MMQ cells were studied in vitro. PPARα was expressed in a majority of PA. PPARα immunostaining was observed in 93.7% PRL-PA vs. 60.6% NFPA (p=0.016), the opposite being found for AIP (83.3% in NFPA vs. 43.7% in PRL-PA, p=0.003). PPARα expression was unrelated to gonadotroph differentiation in NFPA, but positively correlated with tumour volume in PRL-PA. Both drugs significantly reduced MMQ cell growth at high concentrations (100-200 μM). At the same time, despite modest stimulating effects on PRL secretion were observed, these were overcome by the reduction in cell number. In conclusion, PPARα is commonly expressed by PRL-PA and NFPA, regardless of AIP, and may represent a new target of PPARα agonists.

Non-conventional role of haemoglobin beta in breast malignancy

Background:Besides its role as oxygen transporter, recent findings suggest that haemoglobin beta (HBB) may have roles in other contexts.Methods:We evaluated the impact of HBB expression in primary human breast cancers, and in breast cancer cell lines overexpressing HBB by in vitro and in vivo studies. Publicly available microarray databases were used to perform multivariate survival analyses.Results:A significantly higher expression of HBB was observed in invasive carcinoma histotypes vs in situ counterparts, along with a positive correlation between HBB and the Ki67 proliferation marker. HBB-overexpressing breast cancer cells migrate and invade more, show HIF-1α upregulation and their conditioned media enhances angiogenesis. Blocking the oxygen-binding site of HBB reverts the increase of migration and HIF-1α upregulation observed in HBB-overexpressing breast cancer cells. Orthotopically implanted MDA-MB-231 overexpressing HBB (MDA-HBB) generated tumours with faster growth rate and increased neoangiogenesis. Moreover, local recurrence and visceral metastases were observed only in MDA-HBB-implanted mice. Similar results were observed with 4T1 mouse breast cancer cells. Finally, bioinformatics analyses of public data sets correlated high HBB expression with lower overall survival.Conclusions:HBB expression increases breast cancer cells aggressiveness and associates with poor prognosis, pointing to HBB as a novel biomarker for breast cancer progression.

An “Expressionist” Portrait of Mummy Liver

Despite marked postmortem changes, internal organs are often well preserved in naturally mummified bodies. With some obvious limitations and particular attention, histologic examination of such specimens provides useful information on our ancestors’ diseases.The liver is essentially an epithelial organ with a fibrovascular framework. After death, the prompt autolytic changes rapidly delete hepatic cells, but the fibrous portal skeleton of the liver commonly persists. Whereas antemortem patterns of epithelial cells are often destroyed, the conditions characterized by portal alterations may remain recognizable. For this reason, the diagnosis of hepatitis in desiccated bodies cannot be made, and only the different forms of cirrhosis may be identified reliably. The liver of a natural mummy, belonging to a woman aged 50 ± 3 years who lived in the 19th century in Goriano Valli (L’Aquila, central Italy), did not show any sign of hepatic disease but, surprisingly, demonstrated the rules of expressionism long before Pablo Picasso (Figure 1). In particular, the microscopic picture suggested a face depicted with jagged and nervous lines, recalling the works of Egon Schiele (1890-1918), a key figure of the “Viennese Secession.” His own tableaux, mostly portraits of great psychologic intensity, display a tormented vision of life and transmit a strange and fascinating sense of perturbation. Human histopaleopathology not only shows morphologic characters of ancient tissues, but sometimes offers interesting remarks about fine arts. References