The Multifaceted Functions of Prion Protein (PrPC) in Cancer.

The cellular prion protein (PrPC) is a glycoprotein anchored to the cell surface by glycosylphosphatidylinositol (GPI). PrPC is expressed both in the brain and in peripheral tissues. Investigations on PrPC's functions revealed its direct involvement in neurodegenerative and prion diseases, as well as in various physiological processes such as anti-oxidative functions, copper homeostasis, trans-membrane signaling, and cell adhesion. Recent findings have revealed the ectopic expression of PrPC in various cancers including gastric, melanoma, breast, colorectal, pancreatic, as well as rare cancers, where PrPC promotes cellular migration and invasion, tumor growth, and metastasis. Through its downstream signaling, PrPC has also been reported to be involved in resistance to chemotherapy and tumor cell apoptosis. This review summarizes the variance of expression of PrPC in different types of cancers and discusses its roles in their development and progression, as well as its use as a potential target to treat such cancers.

Effects of Prokineticins on Cerebral Cell Function and Blood-Brain Barrier Permeability.

Prokineticins are a family of small proteins with diverse roles in various tissues, including the brain. However, their specific effects on different cerebral cell types and blood-brain barrier (BBB) function remain unclear. The aim of this study was to investigate the effects of PROK1 and PROK2 on murine cerebral cell lines, bEnd.3, C8.D30, and N2a, corresponding to microvascular endothelial cells, astrocytes and neurons, respectively, and on an established BBB co-culture model. Western blot analysis showed that prokineticin receptors (PROKR1 and PROKR2) were differentially expressed in the considered cell lines. The effect of PROK1 and PROK2 on cell proliferation and migration were assessed using time-lapse microscopy. PROK1 decreased neural cells' proliferation, while it had no effect on the proliferation of endothelial cells and astrocytes. In contrast, PROK2 reduced the proliferation of all cell lines tested. Both PROK1 and PROK2 increased the migration of all cell lines. Blocking PROKRs with the PROKR1 antagonist (PC7) and the PROKR2 antagonist (PKR-A) inhibited astrocyte PROK2-mediated migration. Using the insert co-culture model of BBB, we demonstrated that PROKs increased BBB permeability, which could be prevented by PROKRs' antagonists.

Role of NLRP7 in Normal and Malignant Trophoblast Cells.

Gestational choriocarcinoma (CC) is an aggressive cancer that develops upon the occurrence of abnormal pregnancies such as Hydatidiform moles (HMs) or upon non-molar pregnancies. CC cells often metastasize in multiple organs and can cause maternal death. Recent studies have established an association between recurrent HMs and mutations in the Nlrp7 gene. NLRP7 is a member of a new family of proteins that contributes to innate immune processes. Depending on its level of expression, NLRP7 can function in an inflammasome-dependent or independent pathway. To date, the role of NLRP7 in normal and in malignant human placentation remains to be elucidated. We have recently demonstrated that NLRP7 is overexpressed in CC trophoblast cells and may contribute to their acquisition of immune tolerance via the regulation of key immune tolerance-associated factors, namely HLA family, ßCG and PD-L1. We have also demonstrated that NLRP7 increases trophoblast proliferation and decreases their differentiation, both in normal and tumor conditions. Actual findings suggest that NLRP7 expression may ensure a strong tolerance of the trophoblast by the maternal immune system during normal pregnancy and may directly affect the behavior and aggressiveness of malignant trophoblast cells. The proposed review summarizes recent advances in the understanding of the significance of NLRP7 overexpression in CC and discusses its multifaceted roles, including its function in an inflammasome-dependent or independent pathways.

Evidence-Based View of Safety and Effectiveness of Prokineticin Receptors Antagonists during Pregnancy.

Endocrine gland derived vascular endothelial growth factor (EG-VEGF) is a canonical member of the prokineticin (PROKs) family. It acts via the two G-protein coupled receptors, namely PROKR1 and PROKR2. We have recently demonstrated that EG-VEGF is highly expressed in the human placenta; contributes to placental vascularization and growth and that its aberrant expression is associated with pregnancy pathologies including preeclampsia and fetal growth restriction. These findings strongly suggested that antagonization of its receptors may constitute a potential therapy for the pregnancy pathologies. Two specific antagonists of PROKR1 (PC7) and for PROKR2 (PKRA) were reported to reverse PROKs adverse effects in other systems. In the view of using these antagonists to treat pregnancy pathologies, a proof of concept study was designed to determine the biological significances of PC7 and PKRA in normal pregnancy outcome. PC7 and PKRA were tested independently or in combination in trophoblast cells and during early gestation in the gravid mouse. Both independent and combined treatments uncovered endogenous functions of EG-VEGF. The independent use of antagonists distinctively identified PROKR1 and PROKR2-mediated EG-VEGF signaling on trophoblast differentiation and invasion; thereby enhancing feto-placental growth and pregnancy outcome. Thus, our study provides evidence for the potential safe use of PC7 or PKRA to improve pregnancy outcome.

Development of Conformational Antibodies to Detect Bcl-xL's Amyloid Aggregates in Metal-Induced Apoptotic Neuroblastoma Cells.

Bcl-xL, a member of the Bcl-2 family, is a pro-survival protein involved in apoptosis regulation. We have previously reported the ability of Bcl-xL to form various types of fibers, from native to amyloid conformations. Here, we have mimicked the effect of apoptosis-induced caspase activity on Bcl-xL by limited proteolysis using trypsin. We show that cleaved Bcl-xL (ΔN-Bcl-xL) forms fibers that exhibit the features of amyloid structures (BclxLcf37). Moreover, three monoclonal antibodies (mAbs), produced by mouse immunization and directed against ΔN-Bcl-xL or Bcl-xL fibers, were selected and characterized. Our results show that these mAbs specifically target ΔN-Bcl-xL in amyloid fibers in vitro. Upon metal-stress-induced apoptosis, these mAbs are able to detect the presence of Bcl-xL in amyloid aggregates in neuroblastoma SH-SY5Y cell lines. In conclusion, these specific mAbs directed against amyloidogenic conformations of Bcl-xL constitute promising tools for studying, in vitro and in cellulo, the contribution of Bcl-xL in apoptosis. These mAbs may further help in developing new diagnostics and therapies, considering Bcl-xL as a strategic target for treating brain lesions relevant to stroke and neurodegenerative diseases.

Magnetic Nanoparticles Applications for Amyloidosis Study and Detection: A Review.

Magnetic nanoparticles (MNPs) have great potential in biomedical and clinical applications because of their many unique properties. This contribution provides an overview of the MNPs mainly used in the field of amyloid diseases. The first part discusses their use in understanding the amyloid mechanisms of fibrillation, with emphasis on their ability to control aggregation of amyloidogenic proteins. The second part deals with the functionalization by various moieties of numerous MNPs' surfaces (molecules, peptides, antibody fragments, or whole antibodies of MNPs) for the detection and the quantification of amyloid aggregates. The last part of this review focuses on the use of MNPs for magnetic-resonance-based amyloid imaging in biomedical fields, with particular attention to the application of gadolinium-based paramagnetic nanoparticles (AGuIX), which have been recently developed. Biocompatible AGuIX nanoparticles show favorable characteristics for in vivo use, such as nanometric and straightforward functionalization. Their properties have enabled their application in MRI. Here, we report that AGuIX nanoparticles grafted with the Pittsburgh compound B can actively target amyloid aggregates in the brain, beyond the blood⁻brain barrier, and remain the first step in observing amyloid plaques in a mouse model of Alzheimer's disease.

Mass and charge distributions of amyloid fibers involved in neurodegenerative diseases: mapping heterogeneity and polymorphism.

Heterogeneity and polymorphism are generic features of amyloid fibers with some important effects on the related disease development. We report here the characterization, by charge detection mass spectrometry, of amyloid fibers made of three polypeptides involved in neurodegenerative diseases: Aß1-42 peptide, tau and α-synuclein. Beside the mass of individual fibers, this technique enables to characterize the heterogeneity and the polymorphism of the population. In the case of Aß1-42 peptide and tau protein, several coexisting species could be distinguished and characterized. In the case of α-synuclein, we show how the polymorphism affects the mass and charge distributions.

Multimodal imaging Gd-nanoparticles functionalized with Pittsburgh compound B or a nanobody for amyloid plaques targeting.

AIM: Gadolinium-based nanoparticles were functionalized with either the Pittsburgh compound B or a nanobody (B10AP) in order to create multimodal tools for an early diagnosis of amyloidoses. MATERIALS & METHODS: The ability of the functionalized nanoparticles to target amyloid fibrils made of ß-amyloid peptide, amylin or Val30Met-mutated transthyretin formed in vitro or from pathological tissues was investigated by a range of spectroscopic and biophysics techniques including fluorescence microscopy. RESULTS: Nanoparticles functionalized by both probes efficiently interacted with the three types of amyloid fibrils, with KD values in 10 micromolar and 10 nanomolar range for, respectively, Pittsburgh compound B and B10AP nanoparticles. Moreover, they allowed the detection of amyloid deposits on pathological tissues. CONCLUSION: Such functionalized nanoparticles could represent promising flexible and multimodal imaging tools for the early diagnostic of amyloid diseases, in other words, Alzheimer's disease, Type 2 diabetes mellitus and the familial amyloidotic polyneuropathy.

Gd-nanoparticles functionalization with specific peptides for ß-amyloid plaques targeting.

BACKGROUND: Amyloidoses are characterized by the extracellular deposition of insoluble fibrillar proteinaceous aggregates highly organized into cross-ß structure and referred to as amyloid fibrils. Nowadays, the diagnosis of these diseases remains tedious and involves multiple examinations while an early and accurate protein typing is crucial for the patients' treatment. Routinely used neuroimaging techniques such as magnetic resonance imaging (MRI) and positron emission tomography (PET) using Pittsburgh compound B, [(11)C]PIB, provide structural information and allow to assess the amyloid burden, respectively, but cannot discriminate between different amyloid deposits. Therefore, the availability of efficient multimodal imaging nanoparticles targeting specific amyloid fibrils would provide a minimally-invasive imaging tool useful for amyloidoses typing and early diagnosis. In the present study, we have functionalized gadolinium-based MRI nanoparticles (AGuIX) with peptides highly specific for Aß amyloid fibrils, LPFFD and KLVFF. The capacity of such nanoparticles grafted with peptide to discriminate among different amyloid proteins, was tested with Aß(1-42) fibrils and with mutated-(V30M) transthyretin (TTR) fibrils. RESULTS: The results of surface plasmon resonance studies showed that both functionalized nanoparticles interact with Aß(1-42) fibrils with equilibrium dissociation constant (Kd) values of 403 and 350 µM respectively, whilst they did not interact with V30M-TTR fibrils. Similar experiments, performed with PIB, displayed an interaction both with Aß(1-42) fibrils and V30M-TTR fibrils, with Kd values of 6 and 10 µM respectively, confirming this agent as a general amyloid fibril marker. Thereafter, the ability of functionalized nanoparticle to target and bind selectively Aß aggregates was further investigated by immunohistochemistry on AD like-neuropathology brain tissue. Pictures clearly indicated that KLVFF-grafted or LPFFD-grafted to AGuIX nanoparticle recognized and bound the Aß amyloid plaque localized in the mouse hippocampus. CONCLUSION: These results constitute a first step for considering these functionalized nanoparticles as a valuable multimodal imaging tool to selectively discriminate and diagnose amyloidoses.

Amyloid fibrils formed by the programmed cell death regulator Bcl-xL.

The accumulation of amyloid fibers due to protein misfolding is associated with numerous human diseases. For example, the formation of amyloid deposits in neurodegenerative pathologies is correlated with abnormal apoptosis. We report here the in vitro formation of various types of aggregates by Bcl-xL, a protein of the Bcl-2 family involved in the regulation of apoptosis. Bcl-xL forms aggregates in three states, micelles, native-like fibrils, and amyloid fibers, and their biophysical characterization has been performed in detail. Bcl-xL remains in its native state within micelles and native-like fibrils, and our results suggest that native-like fibrils are formed by the association of micelles. Formation of amyloid structures, that is, nonnative intermolecular ß-sheets, is favored by the proximity of proteins within fibrils at the expense of the Bcl-xL native structure. Finally, we provide evidence of a direct relationship between the amyloid character of the fibers and the tertiary-structure stability of the native Bcl-xL. The potential causality between the accumulation of Bcl-xL into amyloid deposits and abnormal apoptosis during neurodegenerative diseases is discussed.

Acute ileal inflammatory cytokine response induced by irradiation is modulated by subdiaphragmatic vagotomy.

Neural involvement plays a role in the genesis of the peripheral inflammatory process that contributes to the irradiation intestinal disorders. However, little is known about the role of vagus nerve in modulating inflammatory process in rat. Here, we have shown that the NF-kappaB activation was consistent with the acute overexpression of pro-inflammatory cytokines (IL- 1beta, TNF-alpha, IL-6) at 3, 6, and 12 h induced by whole-body irradiation (8 Gy). Subdiaphragmatic vagotomy reduced NF-kappaB activation and cytokine transcription in the early period post-irradiation. In contrast, vagotomy amplified overexpression of irradiation-induced anti-cytokines (IL-10, IL-1Ra) and of receptors involved in anti-inflammatory effects (IL- 1RII, TNFRII). These results show that the vagus nerve is a pro-inflammatory pathway in early irradiation-induced intestinal inflammation.

Acute induction of inflammatory cytokine expression after gamma-irradiation in the rat: effect of an NF-kappaB inhibitor.

PURPOSE: The pathologic changes within the intestinal muscle layer may be at the origin of the cytokines that account for acute radiation-induced inflammation. We were specifically interested in evaluating the efficacy of an inhibitor of nuclear transcription factor kappa B (NF-kappaB) activation that is involved in regulating cytokine expression. METHODS AND MATERIALS: Cytokine expression was analyzed in the ileal muscularis layer by reverse transcriptase-polymerase chain reaction (RT-PCR) at 3 h, 6 h, and 3 days after a 10-Gy gamma whole-body irradiation of rats. Caffeic acid phenethyl ester (CAPE) was injected intraperitoneally (30 mg/kg) 15 min before irradiation and once a day for 3 days. RESULTS: Interleukin (IL)-1beta, tumor necrosis factor alpha (TNF-alpha), and IL-6 mRNA increased at 3 h and 6 h after irradiation, and expression of IL-6 and IL-8 was elevated at 3 days. On the other hand, levels of the anti-inflammatory cytokine IL-10 were markedly lower on Day 3. Overexpression of IL-6 on Day 3 was combined with upregulation of the IL-6 receptors (gp130/gp80) and suppressor of cytokine signaling-3 (SOCS3) genes. CAPE treatment did not significantly change IL-1beta or TNF-alpha expressions in the irradiated rats; it increased IL-10 expression at 6 h but had no effect on it on Day 3. CAPE treatment inhibited the radiation-induced expression of IL-6, IL-6 receptors (IL-6rs), and SOCS3 at 3 days. CONCLUSION: In vivo, irradiation induced a cascade of inflammatory responses that involved the transcription factor NF-kappaB; this inflammation was reduced by CAPE treatment.

Involvement of primary afferent nerves after abdominal irradiation: consequences on ileal contractile activity and inflammatory mediator release in the rat.

In this study we analyzed the role of substance P (SP) from afferent nerves in ileum contractibility and in the release of inflammatory mediators (neurotensin, Il-1beta, and TNF-alpha) in ileal mucosa and muscularis layers after a 10-Gy gamma-irradiation of the abdomen. Six hours after irradiation, SP concentrations were lower than in control rats, and 3 days after irradiation SP-induced contractile activity was higher. Irradiation significantly increased the levels of neurotensin, Il-1beta, and TNF-alpha in both layers. Pretreatment with capsaicin depleted afferent nerve endings of SP and reduced SP levels by about 50%. Capsaicin treatment reduced SP concentrations further, beyond the levels due to irradiation, thereby suggesting that all sources of SP are affected by irradiation. Capsaicin treatment prevented the irradiation from affecting SP-induced contractile response or increasing neurotensin levels. This finding suggests that SP released by afferent nerve endings controls these functions. Proinflammatory cytokine release was not reduced by capsaicin treatment.