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Gallstone ileus is an uncommon cause of intestinal obstruction. It is caused when a gallstone migrates through an enterobiliary fistula (most often between the duodenum and the gallbladder) and is impacted in the digestive system, most often in the terminal ileum toward the ileocaecal valve., Case Presentation: Here the authors report the case of a 74-year-old woman who was admitted to Compiegne Hospital in France for a gallstone ileus with the sigmoid colon as the impaction site, which is an even more rare cause of intestinal obstruction. The enterobiliary fistula was between the colon and the gallbladder.The gallstone was removed surgically with a colotomy, without treating the fistula, and after a failed endoscopic attempt to extract the stone. The follow-up was without complications, and a colposcopy showed spontaneous closure of the fistula after 6 weeks., Discussion and Conclusion: The surgical closure of an enterobiliary fistula is an option that should be considered, but it can lead to higher morbidity. That is why the authors opted out of it, especially considering that spontaneous closure of the fistulae can happen, as it did in our case., Competing Interests: The authors declare that they have no financial conflict of interest with regard to the content of this report., (© 2023 the Author(s). Published by Wolters Kluwer Health, Inc.)

A pericyte-like differentiation of human adipose-derived mesenchymal stem cells (ASCs) was tested in in vitro experiments for possible therapeutic applications in cases of diabetic retinopathy (DR) to replace irreversibly lost pericytes. For this purpose, pericyte-like ASCs were obtained after their growth in a specific pericyte medium. They were then cultured in high glucose conditions to mimic the altered microenvironment of a diabetic eye. Several parameters were monitored, especially those particularly affected by disease progression: cell proliferation, viability and migration ability; reactive oxygen species (ROS) production; inflammation-related cytokines and angiogenic factors. Overall, encouraging results were obtained. In fact, even after glucose addition, ASCs pre-cultured in the pericyte medium (pmASCs) showed high proliferation rate, viability and migration ability. A considerable increase in mRNA expression levels of the anti-inflammatory cytokines transforming growth factor-β1 (TGF-β1) and interleukin-10 (IL-10) was observed, associated with reduction in ROS production, and mRNA expression of pro-inflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor-α (TNF-α), and angiogenic factors. Finally, a pmASC-induced better organization of tube-like formation by retinal endothelial cells was observed in three-dimensional co-culture. The pericyte-like ASCs obtained in these experiments represent a valuable tool for the treatment of retinal damages occurring in diabetic patients.

Background: Adipose-derived mesenchymal stem cells (ASCs) are characterized by long-term self-renewal and a high proliferation rate. Under adequate conditions, they may differentiate into cells belonging to mesodermal, endodermal or ectodermal lineages. Pericytes support endothelial cells and play an important role in stabilizing the vessel wall at the microcirculation level. The loss of pericytes, as occurs in diabetic retinopathy, results in a breakdown of the blood-retina barrier (BRB) and infiltration of inflammatory cells. In this context, the use of pericyte-like differentiated ASCs may represent a valuable therapeutic strategy for restoring BRB damage., Aim: To test in vitro strategies to obtain pericyte-like differentiation of human ASCs (hASCs)., Methods: Different culture conditions were tested: hASCs cultured in a basal medium supplemented with transforming growth factor β1; and hASCs cultured in a specific pericyte medium (PM-hASCs). In a further sample, pericyte growth supplement was omitted from the PM. In addition, cultures of human retinal pericytes (hRPCs) were used for comparison. Pericyte-like differentiation of hASCs was tested by immunocytochemical staining and western blotting to evaluate the expression of α-smooth muscle actin (α-SMA) and neural/glial antigen 2 (NG2). Interactions between human retinal endothelial cells (hRECs) and different groups of hASCs were investigated in co-culture experiments. In these cases, the expression of typical junctional proteins such as vascular endothelial-Cadherin, zonula occludens-1 and Occludin were assessed in hRECs. In an in vitro model of the BRB, values of trans-endothelial electrical resistance were measured when hRECs were co-cultured with various groups of pretreated hASCs. The values observed were compared with co-cultures of hRECs and hRPCs as well as with cultures of hRECs alone. Three-dimensional co-cultures of hRECs and hRPCs or pericyte-like hASCs in Matrigel were designed to assess their reciprocal localization., Results: After 3-6 d of culture, α-SMA and NG2 immunocytochemistry showed that the closest pericyte-like phenotype was observed when hASCs were cultured in Pericyte Medium (PM-hASCs). In particular, α-SMA immunoreactivity, already visible at the basal level in pericytes and ASCs, was strongly increased only when transforming growth factor was added to the culture medium. NG2 expression, almost undetectable in most conditions, was substantially increased only in PM-hASCs. Immunocytochemical results were confirmed by western blot analysis. The presence of pericyte growth supplement seems to increase NG2 expression rather than α-SMA, in agreement with its role in maintaining pericytes in the proliferative state. In co-culture experiments, immunoreactivity of vascular endothelial-Cadherin, zonula occludens-1 and Occludin was considerably increased in hRECs when hRPCs or PM-hASCs were also present. Supporting results were found by trans-endothelial electrical resistance measurements, gathered at 3 and 6 d of co-culture. The highest resistance values were obtained when hRECs were co-cultured with hRPCs or PM-hASCs. The pericyte-like phenotype of PM-hASCs was also confirmed in three-dimensional co-cultures in Matrigel, where PM-hASCs and hRPCs similarly localized around the tubular formations made by hRECs., Conclusion: PM-hASCs seem able to strengthen the intercellular junctions between hRECs, likely reinforcing the BRB; thus, hASC-based therapeutic approaches may be developed to restore the integrity of retinal microcirculation., Competing Interests: Conflict-of-interest statement: The authors declare no conflict of interest., (©The Author(s) 2020. Published by Baishideng Publishing Group Inc. All rights reserved.)

Early blood retinal barrier (BRB) dysfunction induced by hyperglycemia was related to increased pro-inflammatory activity of phospholipase A2 (PLA2) and the upregulation of vascular endothelial growth factor A (VEGF-A). Here, we tested the role of VEGF-A in high glucose (HG)-induced damage of human retinal endothelial cells (HRECs) mediated by Ca++-dependent (cPLA2) and Ca++-independent (iPLA2) PLA2s. HRECs were treated with normal glucose (5 mM, NG) or high glucose (25 mM, HG) for 48 h with or without the VEGF-trap Aflibercept (Afl, 40 µg/mL), the cPLA2 inhibitor arachidonoyl trifluoromethyl ketone (AACOCF3; 15 µM), the iPLA2 inhibitor bromoenol lactone (BEL; 5 µM), or VEGF-A (80 ng/mL). Both Afl and AACOCF3 prevented HG-induced damage (MTT and LDH release), impairment of angiogenic potential (tube-formation), and expression of VEGF-A mRNA. Furthermore, Afl counteracted HG-induced increase of phospho-ERK and phospho-cPLA2 (immunoblot). VEGF-A in HG-medium increased glucose toxicity, through upregulation of phospho-ERK, phospho-cPLA2, and iPLA2 (about 55%, 45%, and 50%, respectively); immunocytochemistry confirmed the activation of these proteins. cPLA2 knockdown by siRNA entirely prevented cell damage induced by HG or by HG plus VEGF-A, while iPLA2 knockdown produced a milder protective effect. These data indicate that VEGF-A mediates the early glucose-induced damage in retinal endothelium through the involvement of ERK1/2/PLA2 axis activation.

The study of strial pericytes has gained great interest as they are pivotal for the physiology of stria vascularis. To provide an easily accessible in vitro model, here we described a growth medium-based approach to obtain and cultivate primary bovine cochlear pericytes (BCP) from the stria vascularis of explanted bovine cochleae. We obtained high-quality pericytes in 8-10 days with a > 90% purity after the second passage. Immunocytochemical analysis showed a homogeneous population of cells expressing typical pericyte markers, such as neural/glial antigen 2 (NG2), platelet-derived growth factor receptorβ (PDGFRβ), α-smooth muscle actin (α-SMA), and negative for the endothelial marker von Willebrand factor. When challenged with tumor necrosis factor or lipopolysaccharide, BCP changed their shape, similarly to human retinal pericytes (HRPC). The sensitivity of BCP to ototoxic drugs was evaluated by challenging with cisplatin or gentamicin for 48 hr. Compared to human retinal endothelial cells and HRPC, cell viability of BCP was significantly lower ( p < 0.05) after the treatment with gentamicin or cisplatin. These data indicate that our protocol provides a simple and reliable method to obtain highly pure strial BCP. Furthermore, BCP are suitable to assess the safety profile of molecules which supposedly exert ototoxic activity, and may represent a valid alternative to in vivo tests., (© 2018 Wiley Periodicals, Inc.)

When a contaminant interacts with biotic components of a marine ecosystem, it causes a series of changes that can compromise an entire community (Stebbing, 1985). This present study wants to focus on changes in the gills of a bioindicator benthic organism, Coris julis, collected in Milazzo (Messina, Italy), characterized by a strong anthropical impact), compared with individuals from the control site (Marinello, Messina). RT-PCR has been used for both MT and HSP70, and the respective mRNAs have been visualized by FISH. MT and HSP70 expression levels increased in individuals collected in Milazzo. The presence of numerous apoptotic and proliferating cells and the analysis of several neuronal markers by immunohistochemical method give information about the adaptation to a heavy metal mixture. The obtained results show that, in stressed fishes, defensive processes increase to maintain the normal functions of the organs more exposed to the action of polluted substances., (Copyright (c) 2010 Elsevier Inc. All rights reserved.)

Alterations in developmental programming of neuroendocrine and immune system function may critically modulate vulnerability to various diseases. In particular, genetic factors, including gender, may interact with early life events such as exposure to hormones, endotoxins, or neurotoxins, thereby influencing disease predisposition and/or severity, but little is known about the role of the astroglial cell compartment and its mediators in this phenomenon. Indeed, in the context of innate inflammatory mechanisms, a dysfunction of the astroglial cell compartment is believed to contribute to the selective degeneration of dopaminergic (DA) neurons in the substantia nigra pars compacta in Parkinson's disease (PD) and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) model of PD. Hence, in response to brain injury the roles of astrocytes and microglia are very dynamic and cell type-dependent, in that they may exert the known proinflammatory (harmful) effects, but in certain circumstances they can turn into highly protective cells and exert anti-inflammatory (beneficial) functions, thereby facilitating neuronal recovery and repair. Here, we summarize our work suggesting a chief role of hormonal programming of glial response to inflammation and oxidative stress in MPTP-induced loss of DA neuron functionality and demonstrate that endogenous glucocorticoids and the female hormone estrogen (E(2)) inhibit the aberrant neuroinflammatory cascade, protect astrocytes and microglia from programmed cell death, and stimulate recovery of DA neuron functionality, thereby triggering the repair process. The overall results highlight glia as a final common pathway directing neuroprotection versus neurodegeneration. Such recognition of endogenous glial protective pathways may provide a new insight and may contribute to the development of novel therapeutic treatment strategies for PD and possibly other neurodegenerative disorders.

Inflammation and oxidative stress have been closely associated with the pathogenesis of neurodegenerative disorders, including Parkinson's disease (PD). The expression of inducible nitric oxide synthase (iNOS) in astrocytes and microglia and the production of large amounts of nitric oxide (NO) are thought to contribute to dopaminergic neuron demise. Increasing evidence, however, indicates that activated astroglial cells play key roles in neuroprotection and can promote recovery of CNS functions. Endogenous glucocorticoids (GCs) via glucocorticoid receptors (GRs) exert potent anti-inflammatory and immunosuppressive effects and are key players in protecting the brain against stimulation of innate immunity. Here we review our work showing that exposure to a dysfunctional GR from early embryonic life in transgenic (Tg) mice expressing GR antisense RNA represents a key vulnerability factor in the response of nigrostriatal dopaminergic neurons to the neurotoxin, 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), and further report that exacerbation of dopaminergic neurotoxicity with no recovery is determined by failure of astroglia to exert neuroprotective effects. Aberrant iNOS gene expression and increased glia vulnerability to cell death characterized the response of GR-deficient mice to stimulation of innate immunity. More importantly, GR-deficient glial cells failed to protect fetal dopaminergic neurons against oxidative stress-induces cell death, whereas wild-type glia afforded neuroprotection. Thus, lack of iNOS/NO regulation by GCs can program an aberrant GR-NO crosstalk in turn responsible for loss of astroglia neuroprotective function in response to stimulation of innate immunity, pointing to glia and efficient GR-NO dialogue as pivotal factors orchestrating neuroprotection in experimental parkinsonism.

Unconjugated bilirubin (UCB) causes encephalopathy in severely jaundiced neonates by damaging astrocytes and neurons. Astrocytes, which help defend the brain against cytotoxic insults, express the ATP-dependent transporter, multidrug resistance-associated protein 1 (Mrp1), which mediates export of organic anions, probably including UCB. We therefore studied whether exposure to UCB affects the expression and intracellular localization of Mrp1 in cultured mouse astroglial cells (>95% astrocytes). Mrp1 was localized and quantitated by confocal laser scanning microscopy and double immunofluorescence labeling by using specific antibodies against Mrp1 and the astrocyte marker glial fibrillary acidic protein, plus the Golgi marker wheat germ agglutinin (WGA). In unexposed astrocytes, Mrp1 colocalized with WGA in the Golgi apparatus. Exposure to UCB at a low unbound concentration (Bf) of 40 nM caused rapid redistribution of Mrp1 from the Golgi throughout the cytoplasm to the plasma membrane, with a peak 5-fold increase in Mrp1 immunofluorescence intensity from 30 to 120 min. Bf above aqueous saturation produced a similar but aborted response. Exposure to this higher Bf for 16 h markedly decreased Trypan blue exclusion and methylthiazoletetrazoilum activity and increased apoptosis 5-fold by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling assay. These toxic effects were modestly increased by inhibition of Mrp1 activity with 3-([3-(2-[7-chloro-2-quinolinyl]ethenyl)phenyl-(3-dimethylamino-3-oxopropyl)-thio-methyl]thio)propanoic acid (MK571). By contrast, Bf=40 nM caused injury only if Mrp1 activity was inhibited by MK571, which also blocked translocation of Mrp1. Our conclusion is that in astrocytes, UCB up-regulates expression of Mrp1 and promotes its trafficking from the Golgi to the plasma membrane, thus moderating cytotoxicity from UCB, presumably by limiting its intracellular accumulation.

Glucocorticoids (GCs) exert via glucocorticoid receptors (GRs) potent anti-inflammatory and immunosuppressive effects. Emerging evidence indicates that an inflammatory process is involved in dopaminergic nigro-striatal neuronal loss in Parkinson's disease. We here report that the GR deficiency of transgenic (Tg) mice expressing GR antisense RNA from early embryonic life has a dramatic impact in "programming" the vulnerability of dopaminergic neurons to 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The GR deficiency of Tg mice exacerbates MPTP-induced toxicity to dopaminergic neurons, as revealed by both severe loss of tyrosine hydroxylase positive nigral neurons and sharp decreases in striatal levels of dopamine and its metabolites. In addition, the late increase in dopamine oxidative metabolism and ascorbic acid oxidative status in GR-deficient mice was far greater than in wild-type (Wt) mice. Inducible nitric oxide synthase (iNOS) was sharply increased in activated astrocytes, macrophages/microglia of GR-deficient as compared with Wt mice. Moreover, GR-deficient microglia produced three- to fourfold higher nitrite levels than Wt mice; these increases preceded the loss of dopaminergic function and were resistant to GR the inhibitory effect of GC, pointing to peroxynitrites as candidate neurotoxic effectors. The iNOS inhibitor N6-(1-iminoethyl)-L-lysine normalized vulnerability of Tg mice, thus establishing a novel link between genetic impairment of GR function and vulnerability to MPTP.