Similar to the well‐recognized phenotypical heterogeneity of hepatocytes, in situ sublobular variations have recently been detected in the cell structure, fenestration patterns, filtrating ...efficiency, surface glycosylation, scavenger function and pathological responses of the sinusoidal lining endothelium. However, unlike other liver cell populations, until now no endothelial cell subpopulations had been isolated or defined with clarity, much less with sublobular/acinar zone‐related differential properties. On the basis of our previous studies showing that periportal segments of mouse liver sinusoids express a significantly higher number of wheat germ agglutinin–binding sites than do perivenous ones, we used this differential feature for in vitro labeling of the specific sublobular derivation of isolated sinusoidal lining endothelial cells to correlate their original lobular position with other features determined on flow cytometry, centrifugal elutriation, discontinuous arabinogalactan density gradients and electron microscopy. Our results revealed additional heterogeneous properties whose association with high or low wheat germ agglutinin–binding capacity made it possible to define in vitro two dominant endothelial cell subpopulations that appear similar to the differential features in the periportal and perivenous sinusoidal segments. Type 1 endothelial cells had low forward angle light scatter and high integrated side scatter, low cytoplasmic porosity index (12% ± 5%) and high wheat germ agglutinin–binding efficiency (160 ± 35 fluorescence intensity units/cell size); these findings are similar to what was observed in situ in the periportal sinusoidal endothelium. On the other hand, type 2 endothelial cells, with high forward angle light scatter and low integrated side scatter, had a high cytoplasmic porosity index (25% ± 8%) and low wheat germ agglutinin–binding efficiency (60 ± 15 fluorescence intensity units/cell size), findings similar to in situ observations of the perivenous sinusoidal lining endothelium. Moreover, these physical and morphological differences entail different cell sedimentation behaviors: type 1 endothelial cell sedimented at high centrifugal elutriation counterflow rates (23 to 37 ml/min) and high arabinogalactan density gradient levels (10% to 15%), whereas type 2 endothelial cell sedimented at low counterflow rates (18 to 23 ml/min) and low density levels (6% to 10%). The combination of these separation procedures made it possible to isolate a 90%‐enriched type 1 endothelial cell population in the 12% to 15% interphase of the 23 and 37 ml/min elutriation flow rates and a 75%‐enriched type 2 endothelial cell population in the 6% to 10% interphase of the 18 and 23 ml/min flow rates. (HEPATOLOGY 1993;18:328–339).
Using fluorescein isothiocyanate‐conjugated ovalbumin (OVA‐FITC), 125I‐mannan, or 125I‐invertase as specific ligands for the mannose receptor, we have quantified its activity in mouse and rat hepatic ...sinusoidal endothelium (HSE), under both basal conditions and after lipopolysaccharide (LPS) or human recombinant interleukin‐1β (IL‐1β) stimulations. Mouse treatment for 4 hours with 5 μg/kg IL‐1β significantly increased OVA‐FITC uptake by HSE. Ligand uptake exhibited a sublobular compartmentalization: In control mice as well as in IL‐1β‐stimulated mice, the ligand distributed preferentially in the periportal and septal areas; no OVA‐FITC was observed in the perivenous sinusoids. In vitro exposure of mouse HSE to 100 pg/mL LPS or 1 ng/mL IL‐1β for 6 hours significantly (P < .01) increased OVA‐FITC uptake. Blocking IL‐1 receptors in HSE by addition of 100 ng/mL IL‐1 receptor antagonist (IL‐1Ra) before stimulation with LPS or IL‐1β abrogated the increase in mannose receptor‐mediated uptake. In vitro endocytosis assays showed that rat HSE uptake of 125I‐mannan or 125I‐invertase progressively increased with both exposure time and concentration of added IL‐1β. Upregulation of mannose receptor‐mediated uptake in response to IL‐1β or LPS was also blocked by previous addition of IL‐1Ra to rat HSE. Flow cytometric analysis showed a significant HSE heterogeneity in mannose receptor‐mediated endocytosis in response to IL‐1β treatment: type I endothelial cells (EC‐I, defined by their small size and high cytoplasmic density) significantly (P < .01) increased OVA‐FITC uptake compared with type II endothelial cells (EC‐II, defined by their large size and low cytoplasmic density). In addition, the subset of EC‐I contained three times more IL‐1β‐binding cells than the EC‐II subset. Because EC‐I and EC‐II are preferentially located in the periportal and perivenous segments of hepatic sinusoids, respectively, these results suggest that IL‐1β, apart from upregulating mannose receptor activity, contributes to the sublobular compartmentalization of this endothelial cell function.
The growth of cancer cells in inflammatory tissue is often observed. This can be the result of favorable conditions for endothelial cell adherence and/or increased production of local growth factors. ...The role of the proinflammatory cytokine interleukin 1 (IL-1) in the prometastatic and growth-promoting environment of inflammation was studied in vivo, and the mechanism of cytokine action was studied in vitro as well. Systemic inflammation was induced by the intravenous injection of IL- beta 3 or lipopolysaccharide (LPS), and the hepatic metastasizing ability of B16 melanoma (B16) cells following intrasplenic injection was studied. IL-1 receptor blockade was accomplished with the use of the IL-1 receptor antagonist (IL-1Ra). In vitro, IL-1Ra was used to assess the mechanism for prometastasis and growth promotion of cultured hepatic sinusoidal endothelium stimulated with LPS. There was a statistically significant (P<.01) enhancement in the parameters of hepatic metastasis when B16 cells were injected intrasplenicaily either 4 hours after IL-1 injection or 6 or 12 hours after LPS injection. IL-1Ra pretreatment reduced IL-1-induced enhancement of metastasis by 73%-87 and completely inhibited the augmentation of metastasis following LPS injection. In vitro, the adherence of melanoma cells to LPS-treated endothelium increased nearly twofold but was completely abrogated when IL-1Ra was added before LPS. Similar to melanoma adherence, a 2.5-fold increase (P<.05) in functional mannose receptors was observed with LPS treatment but was prevented by the addition of IL-1Ra. IL-1Ra did not affect basal mannose-receptor activity in unstimulated epithelium. Mannose-receptor activity and B16 cell adherence significantly correlated (r = .9) with LPS treatment. Conditioned medium from LPS-stimulated epithelium augmented B16 cell proliferation compared with control conditioned medium (P<.01). Production of B16 cell growth factor(s) was markedly reduced (P<.01) when IL-1Ra was added. These results demonstrate that systemic inflammation induces an enhancement of melanoma cell metastasis and growth by IL-1-dependent mechanisms in vivo. In vitro, the mechanism(s) is consistent with IL-1-mediated increase in expression of mannose receptors and production of tumor cell growth factor(s) from the endothelium. Given the multiple and complex cytokine cascade induced in vivo and in vitro during LPS-induced systemic inflammation, IL-1 plays a strategic role. Since IL-1Ra is without side effects in humans, studies on intraoperative infusion of IL-1Ra during tumor resection may be indicated.
Human sperm protein associated with the nucleus on the X chromosome (SPANX) genes encode a protein family (SPANX-A, -B, -C and -D), whose expression is limited to the testis and spermatozoa in normal ...tissues and various tumour cells. SPANX-A/D proteins have been detected in metastatic melanoma cells, but their contribution to cancer development and the underlying molecular mechanisms of skin tumourigenesis remain unknown. Combining functional and proteomic approaches, the present work describes the presence of SPANX-A/D in primary and metastatic human melanoma cells and how it promotes pro-tumoural processes such as cell proliferation, motility and migration. We provide insights into the molecular features of skin tumourigenesis, describing for the first time a multifunctional role of the SPANX-A/D protein family in nuclear function, energy metabolism and cell survival, considered key hallmarks of cancer. A better comprehension of the SPANX-A/D protein subfamily and its molecular mechanisms will help to describe new aspects of tumour cell biology and develop new therapeutic targets and tumour-directed pharmacological drugs for skin tumours.
Cytokines are regulated in acute and chronic inflammation, including rheumatoid arthritis (RA) and myocardial infarction (MI). However, the dynamic windows within which cytokine activity/inhibition ...is desirable in RA and MI change timely and locally during the disease. Therefore, traditional, static delivery regimens are unlikely to meet the idiosyncrasy of these highly dynamic pathophysiological and individual processes. Responsive delivery systems and biomaterials, sensing surrogate markers of inflammation (i.e., matrix metalloproteinases - MMPs) and answering with drug release, may present drug activity at the right time, manner, and place. This article discusses MMPs as surrogate markers for disease activity in RA and MI to clock drug discharge to MMP concentration profiles from MMP-responsive drug delivery systems and biomaterials.
Originally considered to act as a transcriptional co-factor, Pirin has recently been reported to play a role in tumorigenesis and the malignant progression of many tumors. Here, we have analyzed the ...diagnostic and prognostic value of Pirin expression in the early stages of melanoma, and its role in the biology of melanocytic cells. Pirin expression was analyzed in a total of 314 melanoma biopsies, correlating this feature with the patient's clinical course. Moreover, PIR downregulated primary melanocytes were analyzed by RNA sequencing, and the data obtained were validated in human melanoma cell lines overexpressing PIR by functional assays. The immunohistochemistry multivariate analysis revealed that early melanomas with stronger Pirin expression were more than twice as likely to develop metastases during the follow-up. Transcriptome analysis of PIR downregulated melanocytes showed a dampening of genes involved in the G1/S transition, cell proliferation, and cell migration. In addition, an in silico approach predicted that JARID1B as a potential transcriptional regulator that lies between PIR and its downstream modulated genes, which was corroborated by co-transfection experiments and functional analysis. Together, the data obtained indicated that Pirin could be a useful marker for the metastatic progression of melanoma and that it participates in the proliferation of melanoma cells by regulating the slow-cycling JARID1B gene.
Melanoma is the deadliest form of skin cancer due to its ability to colonize distant sites and initiate metastasis. Although these processes largely depend on the lipid-based cell membrane scaffold, ...our understanding of the melanoma lipid phenotype lags behind most other aspects of this tumor cell. Here, we examined a panel of normal human epidermal and nevus melanocytes and primary and metastatic melanoma cell lines to determine whether distinctive cell-intrinsic lipidomes can discern non-neoplastic from neoplastic melanocytes and define their metastatic potential. Lipidome profiles were obtained by UHPLC-ESI mass-spectrometry, and differences in the signatures were analyzed by multivariate statistical analyses. Significant and highly specific changes in more than 30 lipid species were annotated in the initiation of melanoma, whereas less numerous changes were associated with melanoma progression and the non-malignant transformation of nevus melanocytes. Notably, the "malignancy lipid signature" features marked drops in pivotal membrane lipids, like sphingomyelins, and aberrant elevation of ether-type lipids and phosphatidylglycerol and phosphatidylinositol variants, suggesting a previously undefined remodeling of sphingolipid and glycerophospholipid metabolism. Besides broadening the molecular definition of this neoplasm, the different lipid profiles identified may help improve the clinical diagnosis/prognosis and facilitate therapeutic interventions for cutaneous melanoma.
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