Insights in opiates toxicity: impairment of human vascular mesenchymal stromal cells.

The most common pulmonary findings in opiate-related fatalities are congestion and oedema, as well as acute and/or chronic alveolar haemorrhage, the cause of which is thought to be a damage to the capillary endothelium related to ischemia. Human vascular mesenchymal stromal cells (vMSCs) play a fundamental role in tissue regeneration and repair after endothelial cell injury, and they express opioid receptors. The aim of this study was to assess the effect of in vitro morphine exposure on the physiological activity and maintenance of human vMSCs. vMSCs were obtained from abdominal aorta fragments collected during surgery repair and were exposed to incremental doses (0.1 mM, 0.4 mM, 0.8 mM and 1 mM) of morphine sulphate for 7 days. The effect was investigated through cell viability assessment, proliferation assay, reactive oxygen species (ROS) detection assay, senescence-associated ß-galactosidase assay, senescent-related markers (p21WAF1/CIP1 and p16INK4) and the apoptosis-related marker caspase 3. Moreover, an ultrastructural analysis by transmission electron microscopy and in vitro vascular differentiation were evaluated. Results showed a decrease of the cellular metabolic activity, a pro-oxidant and pro-senescence effect, an increase in intracellular ROS and the activation of the apoptosis signalling, as well as ultrastructural modifications and impairment of vascular differentiation after morphine treatment of vMSC. Although confirmation studies are required on real fatal opiate intoxications, the approach based on morphological and immunofluorescence methodologies may have a high potential also as a useful tool or as a complementary method in forensic pathology. The application of these techniques in the future may lead to the identification of new markers and morphological parameters useful as complementary investigations for drug-related deaths.

Implication of Cellular Senescence in Osteoarthritis: A Study on Equine Synovial Fluid Mesenchymal Stromal Cells.

Osteoarthritis (OA) is described as a chronic degenerative disease characterized by the loss of articular cartilage. Senescence is a natural cellular response to stressors. Beneficial in certain conditions, the accumulation of senescent cells has been implicated in the pathophysiology of many diseases associated with aging. Recently, it has been demonstrated that mesenchymal stem/stromal cells isolated from OA patients contain many senescent cells that inhibit cartilage regeneration. However, the link between cellular senescence in MSCs and OA progression is still debated. In this study, we aim to characterize and compare synovial fluid MSCs (sf-MSCs), isolated from OA joints, with healthy sf-MSCs, investigating the senescence hallmarks and how this state could affect cartilage repair. Sf-MSCs were isolated from tibiotarsal joints of healthy and diseased horses with an established diagnosis of OA with an age ranging from 8 to 14 years. Cells were cultured in vitro and characterized for cell proliferation assay, cell cycle analysis, ROS detection assay, ultrastructure analysis, and the expression of senescent markers. To evaluate the influence of senescence on chondrogenic differentiation, OA sf-MSCs were stimulated in vitro for up to 21 days with chondrogenic factors, and the expression of chondrogenic markers was compared with healthy sf-MSCs. Our findings demonstrated the presence of senescent sf-MSCs in OA joints with impaired chondrogenic differentiation abilities, which could have a potential influence on OA progression.

Cell signaling pathways in autosomal-dominant leukodystrophy (ADLD): the intriguing role of the astrocytes.

Autosomal-dominant leukodystrophy (ADLD) is a rare fatal neurodegenerative disorder with overexpression of the nuclear lamina component, Lamin B1 due to LMNB1 gene duplication or deletions upstream of the gene. The molecular mechanisms responsible for driving the onset and development of this pathology are not clear yet. Vacuolar demyelination seems to be one of the most significant histopathological observations of ADLD. Considering the role of oligodendrocytes, astrocytes, and leukemia inhibitory factor (LIF)-activated signaling pathways in the myelination processes, this work aims to analyze the specific alterations in different cell populations from patients with LMNB1 duplications and engineered cellular models overexpressing Lamin B1 protein. Our results point out, for the first time, that astrocytes may be pivotal in the evolution of the disease. Indeed, cells from ADLD patients and astrocytes overexpressing LMNB1 show severe ultrastructural nuclear alterations, not present in oligodendrocytes overexpressing LMNB1. Moreover, the accumulation of Lamin B1 in astrocytes induces a reduction in LIF and in LIF-Receptor (LIF-R) levels with a consequential decrease in LIF secretion. Therefore, in both our cellular models, Jak/Stat3 and PI3K/Akt axes, downstream of LIF/LIF-R, are downregulated. Significantly, the administration of exogenous LIF can partially reverse the toxic effects induced by Lamin B1 accumulation with differences between astrocytes and oligodendrocytes, highlighting that LMNB1 overexpression drastically affects astrocytic function reducing their fundamental support to oligodendrocytes in the myelination process. In addition, inflammation has also been investigated, showing an increased activation in ADLD patients' cells.

Synovium-derived stromal cell-induced osteoclastogenesis: a potential osteoarthritis trigger.

Purpose: To shed light on the idea that mesenchymal stem/stromal cells (MSCs) recruited in synovium (SM) (i.e. Synovium-Derived Stromal Cells, SDSCs) could be involved in Osteoarthritis (OA) pathophysiology. Attention was also paid to a further stromal cell type with a peculiar ultrastructure called telocytes (TCs), whose role is far from clarified. Methods: In the present in vitro study, we compared SDSCs isolated from healthy and OA subjects in terms of phenotype, morphology and differentiation potential as well as in their capability to activate normal Peripheral Blood Mononuclear Cells (PBMCs). Histological, immunohistochemical and ultrastructural analyses were integrated by qRT-PCR and functional resorbing assays. Results: Our data demonstrated that both SDSC populations stimulated the formation of osteoclasts from PBMCs: the osteoclast-like cells generated by healthy-SDSCs via transwell co-cultures were inactive, while OA-derived SDSCs have a much greater effectiveness. Moreover, the presence of TCs was more evident in cultures obtained from OA subjects and suggests a possible involvement of these cells in OA. Conclusions: Osteoclastogenic differentiation capability of PBMCs from OA subjects, also induced by B synoviocytes has been already documented. Here we hypothesized that SDSCs, generally considered for their regenerative potential in cartilage lesions, have also a role in the onset/maintenance of OA. Clinical relevance: Our observations may represent an interesting opportunity for the development of a holistic approach for OA treatment, that considers the multifaceted capability of MSCs in relation to the environment.

Quercetin Exposure Suppresses the Inflammatory Pathway in Intestinal Organoids from Winnie Mice.

Inflammatory bowel diseases (IBDs) are chronic and relapsing immune disorders that result, or possibly originate, from epithelial barrier defects. Intestinal organoids are a new reliable tool to investigate epithelial response in models of chronic inflammation. We produced organoids from the ulcerative colitis murine model Winnie to explore if the chronic inflammatory features observed in the parental intestine were preserved by the organoids. Furthermore, we investigated if quercetin administration to in vitro cultured organoids could suppress LPS-induced inflammation in wild-type organoids (WT-organoids) and spontaneous inflammation in ulcerative colitis organoids (UC-organoids). Our data demonstrate that small intestinal organoids obtained from Winnie mice retain the chronic intestinal inflammatory features characteristic of the parental tissue. Quercetin administration was able to suppress inflammation both in UC-organoids and in LPS-treated WT-organoids. Altogether, our data demonstrate that UC-organoids are a reliable experimental system for investigating chronic intestinal inflammation and pharmacological responses.

HIF1α protein and mRNA expression as a new marker for post mortem interval estimation in human gingival tissue.

Estimating the post mortem interval (PMI) is still a crucial step in Forensic Pathology. Although several methods are available for assessing the PMI, a precise estimation is still quite unreliable and can be inaccurate. The present study aimed to investigate the immunohistochemical distribution and mRNA expression of hypoxia inducible factor (HIF-1α) in post mortem gingival tissues to establish a correlation between the presence of HIF-1α and the time since death, with the final goal of achieving a more accurate PMI estimation. Samples of gingival tissues were obtained from 10 cadavers at different PMIs (1-3 days, 4-5 days and 8-9 days), and were processed for immunohistochemistry and quantitative reverse transcription-polymerase chain reaction. The results showed a time-dependent correlation of HIF-1α protein and its mRNA with different times since death, which suggests that HIF-1α is a potential marker for PMI estimation. The results showed a high HIF-1α protein signal that was mainly localized in the stratum basale of the oral mucosa in samples collected at a short PMI (1-3 days). It gradually decreased in samples collected at a medium PMI (4-5 days), but it was not detected in samples collected at a long PMI (8-9 days). These results are in agreement with the mRNA data. These data indicate an interesting potential utility of Forensic Anatomy-based techniques, such as immunohistochemistry, as important complementary tools to be used in forensic investigations.

Detecting senescent fate in mesenchymal stem cells: a combined cytofluorimetric and ultrastructural approach.

Senescence can impair the therapeutic potential of stem cells. In this study, senescence-associated morphofunctional changes in periosteum-derived progenitor cells (PDPCs) from old and young individuals were investigated by combining cytofluorimetry, immunohistochemistry, and transmission electron microscopy. Cell cycle analysis demonstrated a large number of G0/G1 phase cells in PDPCs from old subjects and a progressive accumulation of G0/G1 cells during passaging in cultures from young subjects. Cytofluorimetry documented significant changes in light scattering parameters and closely correlated with the ultrastructural features, especially changes in mitochondrial shape and autophagy, which are consistent with the mitochondrial-lysosomal axis theory of ageing. The combined morphological, biofunctional, and ultrastructural approach enhanced the flow cytometric study of PDPC ageing. We speculate that impaired autophagy, documented in replicative senescent and old PDPCs, reflect a switch from quiescence to senescence. Its demonstration in a tissue with limited turnover-like the cambium layer of the periosteum, where reversible quiescence is the normal stem cell state throughout life-adds a new piece to the regenerative medicine jigsaw in an ageing society.

Preparation and Evaluation of a Novel Class of Amphiphilic Amines as Antitumor Agents and Nanocarriers for Bioactive Molecules.

PURPOSE: We describe a novel class of antitumor amphiphilic amines (RCn) based on a tricyclic amine hydrophilic head and a hydrophobic linear alkyl tail of variable length. METHODS: We tested the lead compound, RC16, for cytotoxicity and mechanism of cell death in several cancer cell lines, anti tumor efficacy in mouse tumor models, and ability to encapsulate chemotherapy drugs. RESULTS: These compounds displayed strong cytotoxic activity against cell lines derived from both pediatric and adult cancers. The IC50 of the lead compound, RC16, for normal cells including human keratinocytes, human fibroblasts and human umbilical vein endothelial cells was tenfold higher than for tumor cells. RC16 exhibited significant antitumor effects in vivo using several human xenografts and a metastatic model of murine neuroblastoma by both intravenous and oral administration routes. The amphiphilic character of RC16 triggered a spontaneous molecular self-assembling in water with formation of micelles allowing complexation of Doxorubicin, Etoposide and Paclitaxel. These micelles significantly improved the in vitro antitumor activity of these drugs as the enhancement of their aqueous solubility also improved their biologic availability. CONCLUSIONS: RC16 and related amphiphilic amines may be useful as a novel cancer treatment.

Albumin nanocapsules containing fenretinide: pre-clinical evaluation of cytotoxic activity in experimental models of human non-small cell lung cancer.

The present study deals with the preparation of albumin nanocapsules containing fenretinide and their evaluation in experimental models of human non-small cell lung cancer. These nanocapsules showed enhanced antitumor activity with respect to free fenretinide due to the solubilization effect of albumin on the hydrophobic drug, known to improve bioavailability. The high expression of caveolin-1 on the A549 cell surface further enhanced the antitumor activity of the nanoencapsulated fenretinide. Caveolin-1 favored albumin uptake and improved the efficacy of the fenretinide-loaded albumin nanocapsules, especially in 3-D cultures where the densely packed 3-D structures impaired drug diffusibility and severely reduced the activity of the free drug. The efficacy of the fenretinide albumin nanocapsules was further confirmed in tumor xenograft models of A549 by the significant delay in tumor progression observed with respect to control after intravenous administration of the novel formulation. FROM THE CLINICAL EDITOR: This study describes the preparation of fenretinide containing albumin nanocapsules and their evaluation in experimental models of non-small cell lung cancer, showing enhanced antitumor activity compared to free fenretinide.

Simulating tumor microenvironment: changes in protein expression in an in vitro co-culture system.

BACKGROUND: The role of the microenvironment during the initiation and progression of carcinogenesis is thought to be of critical importance, both for the enhanced understanding of fundamental cancer biology as well as for improving molecular diagnostics and therapeutics. The aim of this study was to establish an in vitro model based on a co-culture of healthy human fibroblasts (HFs) and human osteosarcoma cells (MG-63s) to simulate the microenvironment including tumor and healthy cells. METHODS: The HFs and MG-63s were in vitro co-cultured for a period of time ranging from 24 h to 7 days. Cell morphology and organization were studied using phase contrast microscopy while the expression of Human Cartilage Glycoprotein 39 (YKL-40), Vascular Endothelial Growth Factor (VEGF) and Matrix Metalloprotease 1 (MMP1) was investigated by Real Time PCR and Western Blotting. RESULTS: The results showed a characteristic disposition of tumor and healthy co-cultured cells in columns which are not visible in tumor and healthy cells grown singularly. The expression of YKL-40, VEGF and MMP1 significantly changed in co-cultured cells compared to HFs and MG-63s separately cultured. CONCLUSIONS: We concluded that the tumor microenvironment has an influence on the protein expression of the healthy surrounding tissues and the process of tumorigenicity.

Novel PLA microspheres with hydrophilic and bioadhesive surfaces for the controlled delivery of fenretinide.

Novel polylactide (PLA) microspheres endowed with hydrophilic and bioadhesive surfaces as injectable formulations for the controlled release of fenretinide were prepared, using a novel technique based on the co-precipitation of PLA with gelatin, at the interface of a liquid dispersion formed by the addition of N-methylpyrrolidone containing PLA and dextrin (DX), towards an aqueous solution of gelatin (G). The resulting PLA-G-DX microspheres were compared with others prepared by the same technique using polylactide-co-glycolide (PLGA), with or without DX, and with or without phosphatidylcholine. Of the different systems, the PLA-G-DX microspheres had the best morphological, dimensional and functional characteristics. They had the highest drug loading, and their drug release was the most efficient over time without any burst effect. Their in vitro anti-tumoural activity was strongly enhanced with respect to the pure fenretinide. This paralleled the increased drug concentration inside the cells due to their marked bioadhesion to the tumour cell membranes as indicated by scanning electron microscope images.

Antioxidant and Anti-Melanogenesis Effects of Teucrium chamaedrys L. Cell Suspension Extract and Its Main Phenylethanoid Glycoside in B16-F10 Cells.

Teucrium chamaedrys L. is a typical European-Mediterranean species of the genus Teucrium. Among the phenolic compounds belonging to phenylethanoid glycosides (PGs), teucrioside (TS) is only found in this species, and it was previously demonstrated to be produced by in vitro-elicited cell cultures at levels higher than those found in leaves. However, T. chamaedrys cell suspension extracts (Cell-Ex) and pure TS have not been investigated yet for any biological effects. In this study, we evaluated the antioxidant and anti-melanogenesis activity of both Cell-Ex and TS in B16-F10 mouse melanoma cells. The results showed that Cell-Ex inhibited the reactive oxygen species formation evoked in B16-F10 cells by tert-butyl hydroperoxide and 5 J/cm2 of UVA, as well as the melanin increase stimulated by α-MSH or 20 J/cm2 of UVA. In parallel, a TS concentration equivalent to that present in Cell-Ex recorded the same biological effect profile, suggesting the main contribution of TS to the antioxidant and anti-melanogenic properties of Cell-Ex. Both Cell-Ex and TS also modulated the melanogenesis pathway through their ability to inhibit the tyrosinase activity both in a cell-free system and in B16-F10 cells stimulated by α-MSH. These results support the potential cosmeceutical use of Cell-Ex for protection against photooxidative damage and hyperpigmentation.

Antigene MYCN Silencing by BGA002 Inhibits SCLC Progression Blocking mTOR Pathway and Overcomes Multidrug Resistance.

Small-cell lung cancer (SCLC) is the most aggressive lung cancer type, and is associated with smoking, low survival rate due to high vascularization, metastasis and drug resistance. Alterations in MYC family members are biomarkers of poor prognosis for a large number of SCLC. In particular, MYCN alterations define SCLC cases with immunotherapy failure. MYCN has a highly restricted pattern of expression in normal cells and is an ideal target for cancer therapy but is undruggable by traditional approaches. We propose an innovative approach to MYCN inhibition by an MYCN-specific antigene-PNA oligonucleotide (BGA002)-as a new precision medicine for MYCN-related SCLC. We found that BGA002 profoundly and specifically inhibited MYCN expression in SCLC cells, leading to cell-growth inhibition and apoptosis, while also overcoming multidrug resistance. These effects are driven by mTOR pathway block in concomitance with autophagy reactivation, thus avoiding the side effects of targeting mTOR in healthy cells. Moreover, we identified an MYCN-related SCLC gene signature comprehending CNTFR, DLX5 and TNFAIP3, that was reverted by BGA002. Finally, systemic treatment with BGA002 significantly increased survival in MYCN-amplified SCLC mouse models, including in a multidrug-resistant model in which tumor vascularization was also eliminated. These findings warrant the clinical testing of BGA002 in MYCN-related SCLC.

Novel micelles based on amphiphilic branched PEG as carriers for fenretinide.

This study reports on the preparation and evaluation of amphiphilic macromolecules based on branched polyethylene glycol covalently linked with alkyl hydrocarbon chains. These macromolecules easily dissolved in an aqueous environment, with formation of micellar nanoaggregates endowed with hydrophobic inner cores capable of hosting fenretinide by complexation. The complexes increased fenretinide aqueous solubility, while hindering its release as a free drug in an aqueous environment. Particle size analysis indicated dimensional suitability of the complexes for intravenous administration. Neuroblastoma cell lines (SH-SY5Y and NGP) exhibited increased sensitivity to fenretinide in complex as compared to free drug, associated with higher intracellular concentrations of fenretinide observed after treatment with the complex. Transmission electronic microscopy images revealed endocytosis of the micellar complex. Moreover, fenretinide conversion to its metabolite 4-oxo-fenretinide was delayed in cells treated with the complex, further supporting the hypothesis that fenretinide may be absorbed by micellar transport and exposed to the cytoplasm for conversion to its metabolite only after micelle destabilization.

Morphological study of equine amniotic compartment.

Exfoliative cytology of human amniotic fluid (AF) has been extensively studied since 1940s, but no data exist in equine species. The AF compartment represents the environment in which the foetus grows and matures, and its composition changes, reflecting foetal well-being and development. The aim of this study was to describe for the first time the morphology of equine AF cells and amniotic membrane (AM) with light microscopy (LM) and transmission electron microscopy (TEM). AF was collected at parturition within 5 min after the appearance of the AM with a 60 mL syringe from 34 mares and samples of AM were collected from a subset of 7 mares with normal pregnancy hospitalized for attended parturition. For LM observation, a sample of cytocentrifuged fresh AF was stained with May-Grünwald Giemsa and AM sections were stained with H-E. For TEM observation, AF and AM were fixed, embedded in epoxy resins, then sectioned and stained with uranyl acetate and lead citrate solutions. Nucleated and anucleated squamous cells with basophilic cytoplasm, intensely basophilic cornified cells, polymorphonuclear cells, and clusters of eosinophilic amorphous substance were observed. Cells presumably derived from tracheal epithelium and small round nucleated cells with eosinophilic cytoplasm presumably derived from amniotic or urinary epithelium were occasionally found. Lamellar body-like structures (LBs) were present in some epithelial cells. In AM, epithelial, basal and mesenchymal layers were clearly visible with both techniques as previously described. Epithelial cells had several cytoplasmic vacuolization and microvilli were present on apical surface. The connective tissue presented fibroblasts, mesenchymal and rare polymorphonuclear cells, surrounded by abundant extracellular matrix, with distribution of collagen fibres. This is the first report about equine amniotic compartment description by LM and TEM. As recently reported in human medicine, the AM could be a second potential source of pulmonary surfactant, given the finding of LBs inside the cells which could have the same function as in humans. Further studies in samples collected at different gestational ages could increase the knowledge of AF cells and their modification during pregnancy, as well as a better comprehension of the role of AM as a secondary source of pulmonary surfactant in the horse. The diagnostic evaluation of AF cellular composition in high-risk pregnancies may also be investigated.

Esculetin Provides Neuroprotection against Mutant Huntingtin-Induced Toxicity in Huntington's Disease Models.

Huntington's disease (HD) is a neurodegenerative disorder caused by an abnormal CAG trinucleotide repeat expansion within exon 1 of the huntingtin (HTT) gene. This mutation leads to the production of mutant HTT (mHTT) protein which triggers neuronal death through several mechanisms. Here, we investigated the neuroprotective effects of esculetin (ESC), a bioactive phenolic compound, in an inducible PC12 model and a transgenic Drosophila melanogaster model of HD, both of which express mHTT fragments. ESC partially inhibited the progression of mHTT aggregation and reduced neuronal death through its ability to counteract the oxidative stress and mitochondria impairment elicited by mHTT in the PC12 model. The ability of ESC to counteract neuronal death was also confirmed in the transgenic Drosophila model. Although ESC did not modify the lifespan of the transgenic Drosophila, it still seemed to have a positive impact on the HD phenotype of this model. Based on our findings, ESC may be further studied as a potential neuroprotective agent in a rodent transgenic model of HD.

Cellular senescence in vascular wall mesenchymal stromal cells, a possible contribution to the development of aortic aneurysm.

Cellular senescence is a hallmark of ageing and it plays a key role in the development of age-related diseases. Abdominal aortic aneurysm (AAA) is an age related degenerative vascular disorder, characterized by a progressive dilatation of the vascular wall and high risk of rupture over time. Nowadays, no pharmacological therapies are available and the understanding of the molecular mechanisms that lead to AAA onset and development are poorly defined. In this study we investigated the cellular features of senescence in vascular mesenchymal stromal cells, isolated from pathological (AAA - MSCs) and healthy (h - MSCs) segments of human abdominal aorta and their implication in impairing the vascular repair ability of MSCs. Cell proliferation, ROS production, cell surface area, the expression of cyclin dependent kinase inhibitors p21CIP1 and p16INK4a, the activation of the DNA damage response and a dysregulated autophagy showed a senescent state in AAA - MSCs compared to h-MSCs. Moreover, a reduced ability to differentiate toward endothelial cells was observed in AAA - MSCs. All these data suggest that the accumulation of senescent vascular MSCs over time impairs their remodeling ability during ageing. This condition could support the onset and development of AAA.

Assessment of the structural and functional characteristics of human mesenchymal stem cells associated with a prolonged exposure of morphine.

The discovery of the expression of opioid receptors in the skin and their role in orchestrating the process of tissue repair gave rise to questions regarding the potential effects of clinical morphine treatment in wound healing. Although short term treatment was reported to improve tissue regeneration, in vivo chronic administration was associated to an impairment of the physiological healing process and systemic fibrosis. Human mesenchymal stem cells (hMSCs) play a fundamental role in tissue regeneration. In this regard, acute morphine exposition was recently reported to impact negatively on the functional characteristics of hMSCs, but little is currently known about its long-term effects. To determine how a prolonged treatment could impair their functional characteristics, we exposed hMSCs to increasing morphine concentrations respectively for nine and eighteen days, evaluating in particular the fibrogenic potential exerted by the long-term exposition. Our results showed a time dependent cell viability decline, and conditions compatible with a cellular senescent state. Ultrastructural and protein expression analysis were indicative of increased autophagy, suggesting a relation to a detoxification activity. In addition, the enhanced transcription observed for the genes involved in the synthesis and regulation of type I collagen suggested the possibility that a prolonged morphine treatment might exert its fibrotic potential risk, even involving the hMSCs.

Biosynthesis and physico-chemical characterization of high performing peptide hydrogels@graphene oxide composites.

Hydrogels based on short peptide molecules are interesting biomaterials with wide present and prospective use in biotechnologies. A well-known possible drawback of these materials can be their limited mechanical performance. In order to overcome this problem, we prepared Fmoc-Phe3self-assembling peptides by a biocatalytic approach, and we reinforced the hydrogel with graphene oxide nanosheets. The formulation here proposed confers to the hydrogel additional physicochemical properties without hampering peptide self-assembly. We investigated in depth the effect of nanocarbon morphology on hydrogel properties (i.e. morphology, viscoelastic properties, stiffness, resistance to an applied stress). In view of further developments towards possible clinical applications, we have preliminarily tested the biocompatibility of the composites. Our results showed that the innovative hydrogel composite formulation based on FmocPhe3 and GO is a biomaterial with improved mechanical properties that appears suitable for the development of biotechnological applications.

The diagnosis of the cause of the death of Venerina.

Venerina (little Venus) is the name given to a wax model representing a pregnant young woman that was created in Florence (Italy) by Clemente Susini (1754-1814) in 1782. It is currently located in the historic Science Museum of the University of Bologna. The model was constructed so as to enable removal of the thoracic and abdominal walls and various organs, exposing the heart, diaphragm and an opened uterus with a well-developed fetus. The woman is small, about 145 cm (4' 9') tall and of delicate build; she looks like a teenage girl. We know that Clemente Susini worked directly with the cadaver and copied the anatomical preparation exactly. This artist often represented the true structure using a wax mould; the existence of two other versions of this specimen suggests that this model was made in this way. Therefore, Venerina's body may be a faithful representation of a young woman who died while pregnant. Observation of the body confirms that the organs are normal, except for the heart and great vessels. The walls of both ventricles are of equal thickness and the ventricles themselves of approximately equal size. The arch of the aorta and the enlarged pulmonary trunk are connected by a short duct about 3.5 mm in diameter. If this structure represents an open arterial duct, we can deduce that the two ventricles worked under the same conditions of blood pressure, hence their equal wall thickness. If the young woman died from this congenital disease, the cause of death has been diagnosed on a wax model of her body after more than two centuries.