A Preclinical Model of Cutaneous Melanoma Based on Reconstructed Human Epidermis

Malignant melanoma is among the tumor entities with the highest increase of incidence worldwide. To elucidate melanoma progression and develop new effective therapies, rodent models are commonly used. While these do not adequately reflect human physiology, two-dimensional cell cultures lack crucial elements of the tumor microenvironment. To address this shortcoming, we have developed a melanoma skin equivalent based on an open-source epidermal model. Melanoma cell lines with different driver mutations were incorporated into these models forming distinguishable tumor aggregates within a stratified epidermis. Although barrier properties of the skin equivalents were not affected by incorporation of melanoma cells, their presence resulted in a higher metabolic activity indicated by an increased glucose consumption. Furthermore, we re-isolated single cells from the models to characterize the proliferation state within the respective model. The applicability of our model for tumor therapeutics was demonstrated by treatment with a commonly used v-raf murine sarcoma viral oncogene homolog B (BRAF) inhibitor vemurafenib. This selective BRAF inhibitor successfully reduced tumor growth in the models harboring BRAF-mutated melanoma cells. Hence, our model is a promising tool to investigate melanoma development and as a preclinical model for drug discovery.

Endometriosis in the Mouse: Challenges and Progress Toward a ‘Best Fit’ Murine Model

Endometriosis is a prevalent gynecologic condition associated with pelvic pain and infertility characterized by the implantation and growth of endometrial tissue displaced into the pelvis via retrograde menstruation. The mouse is a molecularly well-annotated and cost-efficient species for modeling human disease in the therapeutic discovery pipeline. However, as a non-menstrual species with a closed tubo-ovarian junction, the mouse poses inherent challenges as a preclinical model for endometriosis research. Over the past three decades, numerous murine models of endometriosis have been described with varying degrees of fidelity in recapitulating the essential pathophysiologic features of the human disease. We conducted a search of the peer-reviewed literature to identify publications describing preclinical research using a murine model of endometriosis. Each model was reviewed according to a panel of ideal model parameters founded on the current understanding of endometriosis pathophysiology. Evaluated parameters included method of transplantation, cycle phase and type of tissue transplanted, recipient immune/ovarian status, iterative schedule of transplantation, and option for longitudinal lesion assessment. Though challenges remain, more recent models have incorporated innovative technical approaches such as in vivo fluorescence imaging and novel hormonal preparations to overcome the unique challenges posed by murine anatomy and physiology. These models offer significant advantages in lesion development and readout toward a high-fidelity mouse model for translational research in endometriosis.

Withdrawal from repeated nicotine vapor exposure increases somatic signs of physical dependence, anxiety-like behavior, and brain reward thresholds in adult male rats.

In recent years, there has been a dramatic increase in nicotine vapor consumption via electronic nicotine delivery systems (i.e., e-cigarettes), particularly in adolescents. While recent work has focused on the health effects of nicotine vapor exposure, its effects on the brain and behavior remain unclear. In this study, we assessed the effects that cessation from repeated nicotine vapor exposure had on behavioral and neuronal measures of withdrawal. For Experiment 1, fifty-six adult male rats were tested for plasma cotinine levels, somatic withdrawal signs, and anxiety-like behavior in the elevated plus maze, immediately following precipitated withdrawal from repeated exposure to 12 or 24 mg/mL nicotine vapor. In Experiment 2, twelve adult male rats were tested for intracranial self-stimulation (ICSS) across 14 days of exposure to 24 mg/mL nicotine vapor and across the 14 days immediately following nicotine exposure. Results revealed that plasma cotinine, somatic signs, anxiety-like behavior, and ICSS stimulation thresholds were all observed to be elevated during withdrawal in the 24 mg/mL nicotine group, when compared to vehicle controls (50/50 vegetable glycerin/propylene glycol). The data suggest that cessation from repeated nicotine vapor exposure using our preclinical model leads to nicotine dependence and withdrawal, and demonstrates that the vapor system described in these experiments is a viable pre-clinical model of e-cigarette use in humans. Further characterization of the mechanisms driving nicotine vapor abuse and dependence is needed to improve policies and educational campaigns related to e-cigarette use.

Tissue engineered vascular grafts transform into autologous neovessels capable of native function and growth

Background Tissue-engineered vascular grafts (TEVGs) have the potential to advance the surgical management of infants and children requiring congenital heart surgery by creating functional vascular conduits with growth capacity. Methods Herein, we used an integrative computational-experimental approach to elucidate the natural history of neovessel formation in a large animal preclinical model; combining an in vitro accelerated degradation study with mechanical testing, large animal implantation studies with in vivo imaging and histology, and data-informed computational growth and remodeling models. Results Our findings demonstrate that the structural integrity of the polymeric scaffold is lost over the first 26 weeks in vivo, while polymeric fragments persist for up to 52 weeks. Our models predict that early neotissue accumulation is driven primarily by inflammatory processes in response to the implanted polymeric scaffold, but that turnover becomes progressively mechano-mediated as the scaffold degrades. Using a lamb model, we confirm that early neotissue formation results primarily from the foreign body reaction induced by the scaffold, resulting in an early period of dynamic remodeling characterized by transient TEVG narrowing. As the scaffold degrades, mechano-mediated neotissue remodeling becomes dominant around 26 weeks. After the scaffold degrades completely, the resulting neovessel undergoes growth and remodeling that mimicks native vessel behavior, including biological growth capacity, further supported by fluid–structure interaction simulations providing detailed hemodynamic and wall stress information. Conclusions These findings provide insights into TEVG remodeling, and have important implications for clinical use and future development of TEVGs for children with congenital heart disease.

The Beneficial Effects of Heme Oxygenase 1 and Hydrogen Sulfide Activation in the Management of Neuropathic Pain, Anxiety- and Depressive-like Effects of Paclitaxel in Mice

Chemotherapy-induced peripheral neuropathy constitutes an unresolved clinical problem that severely decreases the quality of the patient’s life. It is characterized by somatosensory alterations, including chronic pain, and a high risk of suffering mental disorders such as depression and anxiety. Unfortunately, an effective treatment for this neuropathology is yet to be found. We investigated the therapeutic potential of cobalt protoporphyrin IX (CoPP), a heme oxygenase 1 inducer, and morpholin-4-ium 4-methoxyphenyl(morpholino) phosphinodithioate dichloromethane complex (GYY4137), a slow hydrogen sulfide (H2S) donor, in a preclinical model of paclitaxel (PTX)-induced peripheral neuropathy (PIPN) in mice. At three weeks after PTX injection, we evaluated the effects of the repetitive administration of 5 mg/kg of CoPP and 35 mg/kg of GYY4137 on PTX-induced nociceptive symptoms (mechanical and cold allodynia) and on the associated emotional disturbances (anxiety- and depressive-like behaviors). We also studied the mechanisms that could mediate their therapeutic properties by evaluating the expression of key proteins implicated in the development of nociception, oxidative stress, microglial activation, and apoptosis in prefrontal cortex (PFC) and dorsal root ganglia (DRG) of mice with PIPN. Results demonstrate that CoPP and GYY4137 treatments inhibited both the nociceptive symptomatology and the derived emotional alterations. These actions were mainly mediated through potentiation of antioxidant responses and inhibiting oxidative stress in the DRG and/or PFC of mice with PIPN. Both treatments normalized some plasticity changes and apoptotic reactions, and GYY4137 blocked microglial activation induced by PTX in PFC. In conclusion, this study proposes CoPP and GYY4137 as good candidates for treating neuropathic pain, anxiety- and depressive-like effects of PTX.

The Route of Vaccine Administration Determines Whether Blood Neutrophils Undergo Long-Term Phenotypic Modifications

Innate immunity modulates adaptive immunity and defines the magnitude, quality, and longevity of antigen-specific T- and B- cell immune memory. Various vaccine and administration factors influence the immune response to vaccination, including the route of vaccine delivery. We studied the dynamics of innate cell responses in blood using a preclinical model of non-human primates immunized with a live attenuated vaccinia virus, a recombinant Modified vaccinia virus Ankara (MVA) expressing a gag-pol-nef fusion of HIV-1, and mass cytometry. We previously showed that it induces a strong, early, and transient innate response, but also late phenotypic modifications of blood myeloid cells after two months when injected subcutaneously. Here, we show that the early innate effector cell responses and plasma inflammatory cytokine profiles differ between subcutaneous and intradermal vaccine injection. Additionally, we show that the intradermal administration fails to induce more highly activated/mature neutrophils long after immunization, in contrast to subcutaneous administration. Different batches of antibodies, staining protocols and generations of mass cytometers were used to generate the two datasets. Mass cytometry data were analyzed in parallel using the same analytical pipeline based on three successive clustering steps, including SPADE, and categorical heatmaps were compared using the Manhattan distance to measure the similarity between cell cluster phenotypes. Overall, we show that the vaccine per se is not sufficient for the late phenotypic modifications of innate myeloid cells, which are evocative of innate immune training. Its route of administration is also crucial, likely by influencing the early innate response, and systemic inflammation, and vaccine biodistribution.

Astrocyte-Derived Pleiotrophin Mitigates Late-Stage Autoimmune CNS Inflammation

Astrocytes are the most abundant glial cells in the central nervous system (CNS) with the capacity to sense and react to injury and inflammatory events. While it has been widely documented that astrocytes can exert tissue-degenerative functions, less is known about their protective and disease-limiting roles. Here, we report the upregulation of pleiotrophin (PTN) by mouse and human astrocytes in multiple sclerosis (MS) and its preclinical model experimental autoimmune encephalomyelitis (EAE). Using CRISPR-Cas9-based genetic perturbation systems, we demonstrate in vivo that astrocyte-derived PTN is critical for the recovery phase of EAE and limits chronic CNS inflammation. PTN reduces pro-inflammatory signaling in astrocytes and microglia and promotes neuronal survival following inflammatory challenge. Finally, we show that intranasal administration of PTN during the late phase of EAE successfully reduces disease severity, making it a potential therapeutic candidate for the treatment of progressive MS, for which existing therapies are limited.