Assessment of the morphology and morphometry of the caudate lobe of the liver using computed tomography.

OBJECTIVE: The caudate lobe differs from the rest of the liver. This study was planned to evaluate the caudate lobe's morphology, morphometry, and vascular structures using computed tomography (CT). PATIENTS AND METHODS: Three hundred eighty-eight cases were evaluated retrospectively in terms of caudate lobe morphology, morphometry, and vascular anatomy from patients undergoing contrast-enhanced abdominal CT for any reason between September 2018 and December 2019. After the application of exclusion criteria, 196 patients were eventually included in the study. RESULTS: One hundred seventeen (59.7%) of the 196 patients were men. The patients' mean age was 57.88 years (ranging from 18 to 82). Morphologically, the caudate lobe was classified as rectangular, piriform, or irregular-shaped, with 117 cases being evaluated as piriform (59.7%), 51 as irregular-shaped (26%), and 28 as rectangular (14.3%). The caudate process was visible in most cases (92.9%). No papillary process was observed in the great majority of patients (87.2%). CONCLUSIONS: Caudate lobe evaluation criteria can be obtained using CT in vivo based on morphological and morphometric values for the caudate lobes yielded by cadaver studies.

Investigation of the effects of swimming exercises in rats given acrylamide.

BACKGROUND: Acrylamide is a toxic substance used in industrial and laboratory processes. Acrylamide exposure has a toxic effect on many systems. Protective mechanisms should be developed against the effects caused by acrylamide. OBJECTIVE: In our study, we investigated whether exercise has a protective effect against the changes that acrylamide will cause in pancreas. METHODS: 32 adult Sprague-Dawley male rats were used. Control group was given only saline. Exercise group was applied swimming exercise for 1hour daily for 4 weeks. Acrylamide group was given 50mg/kg acrylamide by gavage for 4 weeks. Acrylamide+exercise group was applied 50mg/kg acrylamide for 4 weeks and swimming exercise for 1hour daily. After the experiment, fasting blood glucose and oral glucose tolerance test measurements were performed. Then, blood and pancreas samples were taken. RESULTS: Acrylamide exposure caused an increase in fasting blood glucose and oral glucose tolerance, a decrease in insulin levels and oxidative stress in acrylamide group. In exercise group, these values were similar to control group and no significant change was observed in acrylamide+exercise group. While there was an increase in the number of alpha cells in acrylamide group compared to the other groups, here was a decrease in the number of beta cells compared to control group. CONCLUSION: We can say that acrylamide causes changes in the islets of Langerhans by affecting alpha and beta cell numbers. The protective effect of exercise on beta and alpha cell mass was not statistically significant in the acrylamide+exercise group. When the results were examined, the decrease in oxidative stress and the higher number of beta and alpha cells in the acrylamide+exercise group compared to the acrylamide group suggested that 4 weeks of swimming exercise may have an effect on acrylamide exposure.

Extracellular ATP attenuates ischemia-induced caspase-3 cleavage in human endothelial cells.

BACKGROUND: Apoptotic death of endothelial cells (EC) plays a crucial role for the development of ischemic injury. In the present study we investigated the impact of extracellular Adenosine-5'-triphosphate (ATP), either released from cells or exogenously added, on ischemia-induced apoptosis of human EC. METHODS AND RESULTS: To simulate ischemic conditions, cultured human umbilical vein endothelial cells (HUVEC) were exposed to 2 h of hypoxia (Po(2)<4mm Hg) in serum-free medium. Ischemia led to a 1.7-fold (+/-0.4; P<0.05) increase in EC apoptosis compared to normoxic controls as assessed by immunoblotting and immunocytochemistry of cleaved caspase-3. Ischemia-induced apoptosis was accompanied by a 2.3-fold (+/-0.5; P<0.05) increase of extracellular ATP detected by using a luciferin/luciferase assay. Addition of the soluble ecto-ATPase apyrase, enhancing ATP degradation, increased ischemia-induced caspase-3 cleavage. Correspondingly, inhibition of ATP breakdown by addition of the selective ecto-ATPase inhibitor ARL67156 significantly reduced ischemia-induced apoptosis. Extracellular ATP acts on membrane-bound P2Y- and P2X-receptors to induce intracellular signaling. Both, ATP and the P2Y-receptor agonist UTP significantly reduced ischemia-induced apoptosis in an equipotent manner, whereas the P2X-receptor agonist αß-me-ATP did not alter caspase-3 cleavage. The anti-apoptotic effects of ARL67156 and UTP were abrogated when P2-receptors were blocked by Suramin or PPADS. Furthermore, extracellular ATP led to an activation of MEK/ERK- and PI3K/Akt-signaling pathways. Accordingly, inhibition of MEK/ERK-signaling by UO126 or inhibition of PI3K/Akt-signaling by LY294002 abolished the anti-apoptotic effects of ATP. CONCLUSION: The data of the present study indicate that extracellular ATP counteracts ischemia-induced apoptosis of human EC by activating a P2Y-receptor-mediated signaling reducing caspase-3 cleavage.

Intermedin (adrenomedullin2) stabilizes the endothelial barrier and antagonizes thrombin-induced barrier failure in endothelial cell monolayers.

BACKGROUND AND PURPOSE: Intermedin is a member of the calcitonin gene-related-peptide (CGRP) family expressed in endothelial cells and acts via calcitonin receptor-like receptors (CLRs). Here we have analysed the receptors for intermedin and its effect on the endothelial barrier in monolayers of human umbilical vein endothelial cells (HUVECs). EXPERIMENTAL APPROACH: We analysed the effect of intermedin on albumin permeability, contractile machinery, actin cytoskeleton and VE-cadherin in cultured HUVECs. KEY RESULTS: Intermedin concentration-dependently reduced basal endothelial permeability to albumin and antagonized thrombin-induced hyperpermeability. Intermedin was less potent (EC(50) 1.29 ± 0.12 nM) than adrenomedullin (EC(50) 0.24 ± 0.07 nM) in reducing endothelial permeability. These intermedin effects were inhibited by AM(22-52) and higher concentrations of αCGRP(8-37), with pA(2) values of αCGRP(8-37) of 6.4 for both intermedin and adrenomedullin. PCR data showed that HUVEC expressed only the CLR/RAMP2 receptor complex. Intermedin activated cAMP/PKA and cAMP/Epac signalling pathways. Intermedin's effect on permeability was blocked by inhibition of PKA but not of eNOS. Intermedin antagonized thrombin-induced contractile activation, RhoA activation and stress fibre formation. It also induced Rac1 activation, enhanced cell-cell adhesion and antagonized thrombin-induced loss of cell-cell adhesion. Treatment with a specific inhibitor of Rac1 prevented intermedin-mediated barrier stabilization. CONCLUSION AND IMPLICATIONS: Intermedin stabilized endothelial barriers in HUVEC monolayers via CLR/RAMP2 receptors. These effects were mediated via cAMP-mediated inactivation of contractility and strengthening of cell-cell adhesion. These findings identify intermedin as a barrier stabilizing agent and suggest intermedin as a potential treatment for vascular leakage in inflammatory conditions.

Transient hypoxia induces ERK-dependent anti-apoptotic cell survival in endothelial cells.

Ischemia-induced apoptosis of endothelial cells may contribute to tissue injury, organ failure, and transplantation rejection. However, little is known about survival mechanisms capable to counteract endothelial apoptosis. This study investigated the potential role of an endogenous anti-apoptotic response elicited by transient hypoxia, capable to avert ongoing apoptosis in endothelial cells. Experiments were carried out in three different types of cultured endothelial cells (human umbilical vein, pig aorta, and from rat coronary microvasculature). As a pro-apoptotic challenge endothelial cells were cultured in serum-free medium and subjected to hypoxia for 2 h. We found that transient hypoxia reduced caspase 3 activation within 1 h of hypoxia. Accordingly, the number of apoptotic cells was reduced after 24 h of reoxygenation. This was true for all three cell types analyzed. Analysis of Akt and mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) pathways revealed that hypoxia induced a transient activation of ERK 2 but not of Akt. ERK 2 phosphorylation preceded the phosphorylation of pro-apoptotic molecule Bad at Ser112, an inhibitory phosphorylation site specific for ERK. The protective effects of hypoxia regarding Bad phosphorylation, caspase 3 activation, and apoptosis were abolished by MEK 1/2 inhibitors, PD98059 or UO126, as well as by antisense oligonucleotides directed against ERK 1/2. Furthermore, inhibition of this pathway inhibited hypoxia-induced increase in mitochondrial membrane potential. The present study demonstrates that transient hypoxia induces a novel survival mechanism that protects endothelial cells against apoptosis. This endogenous process involves MEK/ERK-mediated inhibition of the pro-apoptotic molecule Bad and caspase 3.

Extracellular ATP induces assembly and activation of the myosin light chain phosphatase complex in endothelial cells.

OBJECTIVES: Extracellular ATP stabilizes the endothelial barrier and inactivates the contractile machinery of endothelial cells. This inactivation relies on dephosphorylation of the regulatory myosin light chain (MLC) due to an activation of the MLC phosphatase (MLCP). To date, activation and function of MLCP in endothelial cells are only partially understood. METHODS: Here, the mechanism of extracellular ATP-mediated activation of MLCP was analyzed in human endothelial cells from umbilical veins. Cells were transfected with the endogenous protein phosphatase 1 (PP1)-specific inhibitor-2 (I-2). RESULTS: Overexpression of I-2 led to inhibition of PP1 activity and abrogation of the ATP-induced dephosphorylation of MLC. This indicates that the PP1 catalytic subunit is the principal phosphatase catalyzing the MLC dephosphorylation induced by extracellular ATP. As demonstrated by immunoprecipitation analysis, extracellular ATP recruits the PP1delta catalytic subunit and the myosin phosphatase targeting subunit (MYPT1) to form a complex. ATP stimulated dephosphorylation of MYPT1 at the inhibitory phosphorylation sites threonine 850 and 696. However, extracellular ATP failed to stimulate MYPT1 dephosphorylation in I-2-overexpressing cells. CONCLUSIONS: The present study shows for the first time that, in endothelial cells, extracellular ATP causes activation of MLCP through recruitment of PP1delta and MYPT1 into a MLCP holoenzyme complex and PP1-mediated reduction of the inhibitory phosphorylation of MYPT1.

Opposite effect of cAMP signaling in endothelial barriers of different origin.

cAMP-mediated signaling mechanisms may destabilize or stabilize the endothelial barrier, depending on the origin of endothelial cells. Here, microvascular coronary [coronary endothelial cells (CEC)] and macrovascular aortic endothelial cell (AEC) monolayers with opposite responses to cAMP were analyzed. Macromolecule permeability, isometric force, activation state of contractile machinery [indicated by phosphorylation of regulatory myosin light chains (MLC), activity of MLC kinase, and MLC phosphatase], and dynamic changes of adhesion complex proteins (translocation of VE-cadherin and paxillin) were determined. cAMP signaling was stimulated by the adenosine receptor agonist 5'-N-(ethylcarboxamido)-adenosine (NECA), the beta-adrenoceptor agonist isoproterenol (Iso), or by the adenylyl cyclase activator forskolin (FSK). Permeability was increased in CEC and decreased in AEC on stimulation with NECA, Iso, or FSK. The effects could be inhibited by the PKA inhibitor Rp-8-CPT-cAMPS and imitated by the PKA activator Sp-cAMPS. Under cAMP/PKA-dependent stimulation, isometric force and MLC phosphorylation were reduced in monolayers of either cell type, due to an activation of MLC phosphatase. In CEC but not in AEC, FSK induced delocalization of VE-cadherin and paxillin from cellular adhesion complexes as indicated by cell fractionation and immunofluorescence microscopy. In conclusion, decline in contractile activation and isometric force contribute to cAMP/PKA-mediated stabilization of barrier function in AEC. In CEC, this stabilizing effect is overruled by cAMP-induced disintegration of cell adhesion structures.

MEK/MAPK as a signaling element in ATP control of endothelial myosin light chain.

Phosphorylation of endothelial myosin light chains (MLC) is a key mechanism in control of endothelial contractile machinery. Extracellular ATP influences endothelial MLC phosphorylation by either activation of Ca(2+)-dependent MLC kinase or Ca(2+)-independent MLC phosphatase. Here, the role of the MEK/MAPK pathway in this signaling was investigated in porcine aortic endothelial cells. Phosphorylation of ERK2 and phosphorylation of MLC were analyzed in cultured aortic endothelial cells. ATP (10 microM) increased ERK2 phosphorylation from basal 17 +/- 3 to 53 +/- 4%, an effect suppressed in the presence of the MEK inhibitors PD-98059 (20 microM) or U0126 (10 microM). Phosphorylation of ERK2 was not dependent on the ATP-induced cytosolic Ca(2+) rise, because it was unaltered when this was suppressed by the Ca(2+) chelator BAPTA (10 microM) or xestospongin C (3 microM), an inhibitor of the inositol 1,4,5-trisphosphate-sensitive Ca(2+) release mechanism of the endoplasmic reticulum. Phosphorylation of ERK2 was neither induced by the adenosine analog 5'-(N-ethylcarboxamido)adenosine (1 microM) nor inhibited in the presence of the adenosine receptor antagonist 8-phenyltheophylline (10 microM). ATP increased MLC kinase activity, and this was blocked in presence of PD-98059. ATP also increased MLC phosphatase activity, which was not inhibited by PD-98059. The MEK/MAPK pathway is a Ca(2+)-independent part of ATP signaling toward MLC kinase but not of ATP signaling toward MLC phosphatase.

ATP reduces macromolecule permeability of endothelial monolayers despite increasing [Ca2+]i.

We investigated the relationship between the ATP-evoked rise of cytosolic Ca2+ concentration ([Ca2+]i) and barrier function in porcine aortic endothelial monolayers. ATP (0.01-100 microM) induced a transient rise of [Ca2+]i and reduced permeability in a concentration-dependent manner. In contrast, the Ca2+ ionophore ionomycin (1 microM) elicited a rise in [Ca2+]i comparable to that induced by ATP (10 microM), but it increased permeability. For the reduction of permeability, nucleotides were found to be in the following order of potency: ATP = ATPgammaS > ADP = UTP. Blockade of adenosine receptors by 8-phenyltheophylline (10 microM) did not affect ATP (10 microM)-induced reduction of permeability. ATP reduced permeability even in endothelial monolayers that had been loaded with the Ca2+ chelator BAPTA to prevent the rise in [Ca2+]i. U-73122 (1 microM), an inhibitor of phospholipase C (PLC), completely abolished the effect of ATP (10 microM) on permeability. It also abolished the translocation of protein kinase C (PKC) in response to ATP, which could also be achieved by the PKC inhibitors Gö-6976 (100 nM) or bisindolylmaleimide I (1 microM). In the presence of PKC inhibitors, however, the permeability effect of ATP was not affected. The presence of inhibitors of adenylate or guanylate cyclase (50 microM SQ-22536 or 20 microM ODQ) prevented changes in cyclic nucleotides but did not affect the permeability effects of ATP. The study shows that ATP reduces macromolecule permeability via a PLC-mediated mechanism that is independent of the concomitant effects of ATP on cytosolic Ca2+, cyclic nucleotides, or PKC.