Zn2+, Mg2+, and Ca2+ are essential minerals required for a plethora of metabolic processes and signaling pathways. Different categories of cation-selective channels and transporters are therefore ...required to tightly control the cellular levels of individual metals in a cell-specific manner. However, the mechanisms responsible for the organismal balance of these essential minerals are poorly understood. Herein, we identify a central and indispensable role of the channel-kinase TRPM7 for organismal mineral homeostasis. The function of TRPM7 was assessed by single-channel analysis of TRPM7, phenotyping of TRPM7-deficient cells in conjunction with metabolic profiling of mice carrying kidney- and intestine-restricted null mutations in Trpm7 and animals with a global “kinase-dead” point mutation in the gene. The TRPM7 channel reconstituted in lipid bilayers displayed a similar permeability to Zn2+ and Mg2+. Consistently, we found that endogenous TRPM7 regulates the total content of Zn2+ andMg2+ in cultured cells. Unexpectedly, genetic inactivation of intestinal rather than kidney TRPM7 caused profound deficiencies specifically of Zn2+, Mg2+, and Ca2+ at the organismal level, a scenario incompatible with early postnatal growth and survival. In contrast, global ablation of TRPM7 kinase activity did not affect mineral homeostasis, reinforcing the importance of the channel activity of TRPM7. Finally, dietary Zn2+ and Mg2+ fortifications significantly extended the survival of offspring lacking intestinal TRPM7. Hence, the organismal balance of divalent cations critically relies on one common gatekeeper, the intestinal TRPM7 channel.
Nonsteroidal anti-inflammatory drugs (NSAIDs) have been proposed as chemopreventive agents for many tumours; however, the mechanism responsible for their anti-neoplastic activity remains elusive and ...the side effects due to cyclooxygenase (COX) inhibition prevent this clinical application.
Molecular biology, in silico, cellular and in vivo tools, including innovative in vivo imaging and classical biochemical assays, were applied to identify and characterise the COX-independent anti-cancer mechanism of NSAIDs.
Here, we show that tumour-protective functions of NSAIDs and exisulind (a sulindac metabolite lacking anti-inflammatory activity) occur through a COX-independent mechanism. We demonstrate these NSAIDs counteract carcinogen-induced proliferation by inhibiting the sirtuin 1 (SIRT1) deacetylase activity, augmenting acetylation and activity of the tumour suppressor p53 and increasing the expression of the antiproliferative gene p21. These properties are shared by all NSAIDs except for ketoprofen lacking anti-cancer properties. The clinical interest of the mechanism identified is underlined by our finding that p53 is activated in mastectomy patients undergoing intraoperative ketorolac, a treatment associated with decreased relapse risk and increased survival.
Our study, for the first-time, links NSAID chemopreventive activity with direct SIRT1 inhibition and activation of the p53/p21 anti-oncogenic pathway, suggesting a novel strategy for the design of tumour-protective drugs.
Immune-mediated graft-versus-tumor (GVT) effects can occur after allogeneic hematopoietic stem cell transplantation (HSCT), but GVT is tightly linked to its main complication, graft-versus-host ...disease (GVHD). Strategies aimed at modulating GVHD, while maintaining the GVT effect, are needed to improve the cure rate of transplant. Given the emerging role of Janus-activated kinase (JAK) signaling in lymphoproliferative and myeloproliferative diseases and its established function at dictating T-cell differentiation, we postulated that JAKs might be potential therapeutic targets through a pharmacologic approach.
We examined the effect of JAK1/JAK2 modulation by ruxolitinib in a mouse model of fully MHC mismatched bone marrow transplant comprising in vivo tumor inoculation.
JAK1/JAK2 inhibition by ruxolitinib improved both overall survival (P = 0.03) and acute GVHD pathologic score at target organs (P ≤ 0.001) of treated mice. In addition, treatment with ruxolitinib was associated with a preserved GVT effect, as evidenced by reduction of tumor burden (P = 0.001) and increase of survival time (P = 0.01). JAK1/JAK2 inhibition did not impair the in vivo acquisition of donor T-cell alloreactivity; this observation may account, at least in part, to the preserved GVT effect. Rather, JAK1/JAK2 inhibition of GVHD was associated with the modulation of chemokine receptor expression, which may have been one factor in the reduced infiltration of donor T cells in GVHD target organs.
These data provide further evidence that JAK inhibition represents a new and potentially clinically relevant approach to GVHD prevention.
Silver nanoparticles (AgNPs) are an important class of nanomaterials used as antimicrobial agents for a wide range of medical and industrial applications. However toxicity of AgNPs and impact of ...their physicochemical characteristics in in vivo models still need to be comprehensively characterized. The aim of this study was to investigate the effect of size and coating on tissue distribution and toxicity of AgNPs after intravenous administration in mice, and compare the results with those obtained after silver acetate administration.
Male CD-1(ICR) mice were intravenously injected with AgNPs of different sizes (10 nm, 40 nm, 100 nm), citrate-or polyvinylpyrrolidone-coated, at a single dose of 10 mg/kg bw. An equivalent dose of silver ions was administered as silver acetate. Mice were euthanized 24 h after the treatment, and silver quantification by ICP-MS and histopathology were performed on spleen, liver, lungs, kidneys, brain, and blood.
For all particle sizes, regardless of their coating, the highest silver concentrations were found in the spleen and liver, followed by lung, kidney, and brain. Silver concentrations were significantly higher in the spleen, lung, kidney, brain, and blood of mice treated with 10 nm AgNPs than those treated with larger particles. Relevant toxic effects (midzonal hepatocellular necrosis, gall bladder hemorrhage) were found in mice treated with 10 nm AgNPs, while in mice treated with 40 nm and 100 nm AgNPs lesions were milder or negligible, respectively. In mice treated with silver acetate, silver concentrations were significantly lower in the spleen and lung, and higher in the kidney than in mice treated with 10 nm AgNPs, and a different target organ of toxicity was identified (kidney).
Administration of the smallest (10 nm) nanoparticles resulted in enhanced silver tissue distribution and overt hepatobiliary toxicity compared to larger ones (40 and 100 nm), while coating had no relevant impact. Distinct patterns of tissue distribution and toxicity were observed after silver acetate administration. It is concluded that if AgNPs become systemically available, they behave differently from ionic silver, exerting distinct and size-dependent effects, strictly related to the nanoparticulate form.
Nutrition science requires more science-based evidences for the development of effective functional diets. To reduce animals for experimental purposes innovative reliable and informative models, ...simulating the complex intestinal physiology, are needed. The aim of this study was to develop a swine duodenum segment perfusion model for the evaluation of nutrient bioaccessibility and functionality across time. At the slaughterhouse, one sow intestine was harvested following Maastricht criteria for organ donation after circulatory death (DCD) for transplantation purposes. Duodenum tract was isolated and perfused in sub-normothermic conditions with heterologous blood after cold ischemia induction. Duodenum segment perfusion model was maintained under controlled pressure conditions through extracorporeal circulation for 3 hours. Blood samples from extracorporeal circulation and luminal content samples were collected at regular intervals for the evaluation of glucose concentration by glucometer, minerals (Na+, Ca2+, Mg2+, K+) by ICP-OES, lactate-dehydrogenase and nitrite oxide by spectrophotometric methods. Dacroscopic observation showed peristaltic activity caused by intrinsic nerves. Glycemia decreased over time (from 44.00±1.20 mg/dL to 27.50±0.41; p < 0.01), suggesting glucose utilization by the tissue confirming the organ viability in line with histological examinations. At the end of the experimental period, intestinal mineral concentrations were lower than their level in blood plasma suggesting their bioaccessibility (p < 0.001). A progressive increase of LDH concentration over time was observed in the luminal content probably related to a loss of viability (from 0.32±0.02 to 1.36±0.02 OD; p < 0.05) confirmed by histological findings that revealed a de-epithelization of the distal portion of duodenum. Isolated swine duodenum perfusion model satisfied the criteria for studying bioaccessibility of nutrients, offering a variety of experimental possibilities in line with 3Rs principle.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Hepatocellular carcinoma (HCC) is a frequent neoplasia and a leading cause of inflammation‐related cancer mortality. Despite that most HCCs arise from persistent inflammatory conditions, pathways ...linking chronic inflammation to cancer development are still incompletely elucidated. We dissected the role of adaptive immunity in the Mdr2 knockout (Mdr2–/–) mouse, a model of inflammation‐associated cancer, in which ablation of adaptive immunity has been induced genetically (Rag2–/–Mdr2–/– and μMt‐Mdr2–/– mice) or with in vivo treatments using lymphocyte‐specific depleting antibodies (anti‐CD20 or anti‐CD4/CD8). We found that activated B and T lymphocytes, secreting fibrogenic tumor necrosis factor alpha (TNFα) and other proinflammatory cytokines, infiltrated liver of the Mdr2–/– mice during chronic fibrosing cholangitis. Lymphocyte ablation, in the Rag2–/–Mdr2–/– and μMt‐Mdr2–/– mice, strongly suppressed hepatic stellate cell (HSC) activation and extracellular matrix deposition, enhancing HSC transition to cellular senescence. Moreover, lack of lymphocytes changed the intrahepatic metabolic/oxidative state, resulting in skewed macrophage polarization toward an anti‐inflammatory M2 phenotype. Remarkably, hepatocarcinogenesis was significantly suppressed in the Rag2–/–Mdr2–/– mice, correlating with reduced TNFα/NF‐κB (nuclear factor kappa B) pathway activation. Ablation of CD20+ B cells, but not of CD4+/CD8+ T cells, in Mdr2–/– mice, promoted senescence‐mediated fibrosis resolution and inhibited the protumorigenic TNFα/NF‐κB pathway. Interestingly, presence of infiltrating B cells correlated with increased tumor aggressiveness and reduced disease‐free survival in human HCC. Conclusion: Adaptive immunity sustains liver fibrosis (LF) and favors HCC growth in chronic injury, by modulating innate components of inflammation and limiting the extent of HSC senescence. Therapies designed for B‐cell targeting may be an effective strategy in LF. (Hepatology 2018;67:1970‐1985).
Atypical chemokine receptors (ACKRs) are regulators of leukocyte traffic, inflammation, and immunity. ACKR2 is a scavenger for most inflammatory CC chemokines and is a negative regulator of ...inflammation. Here we report that ACKR2 is expressed in hematopoietic precursors and downregulated during myeloid differentiation. Genetic inactivation of ACKR2 results in increased levels of inflammatory chemokine receptors and release from the bone marrow of neutrophils with increased anti-metastatic activity. In a model of NeuT-driven primary mammary carcinogenesis ACKR2 deficiency is associated with increased primary tumor growth and protection against metastasis. ACKR2 deficiency results in neutrophil-mediated protection against metastasis in mice orthotopically transplanted with 4T1 mammary carcinoma and intravenously injected with B16F10 melanoma cell lines. Thus, ACKR2 is a key regulator (checkpoint) of mouse myeloid differentiation and function and its targeting unleashes the anti-metastatic activity of neutrophils in mice.
The use of cell-free scaffolds for the regeneration of clinically relevant volumes of soft tissue has been challenged, particularly in the case of synthetic biomaterials, by the difficulty of ...reconciling the manufacturing and biological performance requirements. Here, we investigated in vivo the importance of biomechanical and biochemical cues for conditioning the 3D regenerative microenvironment towards soft tissue formation. In particular, we evaluated the adipogenesis changes related to 3D mechanical properties by creating a gradient of 3D microenvironments with different stiffnesses using 3D Poly(Urethane-Ester-ether) PUEt scaffolds. Our results showed a significant increase in adipose tissue proportions while decreasing the stiffness of the 3D mechanical microenvironment. This mechanical conditioning effect was also compared with biochemical manipulation by loading extracellular matrices (ECMs) with a PPAR-γ activating molecule. Notably, results showed mechanical and biochemical conditioning equivalency in promoting adipose tissue formation in the conditions tested, suggesting that adequate mechanical signaling could be sufficient to boost adipogenesis by influencing tissue remodeling. Overall, this work could open a new avenue in the design of synthetic 3D scaffolds for microenvironment conditioning towards the regeneration of large volumes of soft and adipose tissue, with practical and direct implications in reconstructive and cosmetic surgery.
The melastatin-like transient-receptor-potential-7 protein (TRPM7), harbouring a cation channel and a serine/threonine kinase, has been implicated in thymopoiesis and cytokine expression. Here we ...show, by analysing TRPM7 kinase-dead mutant (Trpm7
) mice, that the enzymatic activity of the receptor is not essential for thymopoiesis, but is required for CD103 transcription and gut-homing of intra-epithelial lymphocytes. Defective T cell gut colonization reduces MHCII expression in intestinal epithelial cells. Mechanistically, TRPM7 kinase activity controls TGF-β-induced CD103 expression and pro-inflammatory T helper 17, but not regulatory T, cell differentiation by modulating SMAD2. Notably, we find that the TRPM7 kinase activity promotes gut colonization by alloreactive T cells in acute graft-versus-host disease. Thus, our results unravel a function of TRPM7 kinase in T cell activity and suggest a therapeutic potential of kinase inhibitors in averting acute graft-versus-host disease.
White striping (WS) is an alteration of breast and thigh muscles of broiler chickens characterized by the presence of white striations parallel to the direction of muscle fibers. This study was ...performed to evaluate the prevalence and the predisposing factors to WS in commercial broilers of different weight reared in northern Italy. Fifty seven broiler flocks, including animals of medium- and heavy-weight, were grossly evaluated at slaughter for the presence of WS. For each flock, breeding data (mean BW at slaughter, ADG, sex, color of skin and fat, genetic line, age, antibiotic treatment, and prevalence of deep pectoral myopathy) were collected and statistically analyzed to assess their correlation with WS. Histology of breast fillets affected by different grades of WS was performed to evaluate potential differences between medium- and heavy-weight broilers. The overall prevalence of WS in medium- and heavy-weight broilers (mean BW 2.59 ± 0.13 kg and 3.64 ± 0.34 kg, respectively) was 70.2 ± 7.9% and 82.51 ± 8.5%, respectively, while the percentage of severe WS was 13.3 ± 7.1% and 25.7 ± 12.8%, respectively. A strong correlation was found between presence of WS, BW at slaughter, and ADG (Pearson correlation = 0.69, P < 0.01; Pearson correlation = 0.67, P < 0.01). WS also closely correlated with the prevalence of deep pectoral myopathy (Spearman's Rho slaughterhouse 1 = 0.74, Spearman's Rho slaughterhouse 2 = 0.51, P < 0.01). No correlation was found between genetics or sanitary status of the flock and WS. Histology confirmed that breasts with WS lesions were affected by a polyphasic degenerative and necrotizing myopathy, and that the lesions, as expected, were more severe in heavy-weight broilers. In conclusion, WS is a major concern in commercial meat poultry reared in Italy, affecting more severely heavier broilers, and it is mainly related to the BW and ADG of animals.
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