Retinal dysfunction in Huntington's disease mouse models concurs with local gliosis and microglia activation.

Huntington's disease (HD) is caused by an aberrant expansion of CAG repeats in the HTT gene that mainly affects basal ganglia. Although striatal dysfunction has been widely studied in HD mouse models, other brain areas can also be relevant to the pathology. In this sense, we have special interest on the retina as this is the most exposed part of the central nervous system that enable health monitoring of patients using noninvasive techniques. To establish the retina as an appropriate tissue for HD studies, we need to correlate the retinal alterations with those in the inner brain, i.e., striatum. We confirmed the malfunction of the transgenic R6/1 retinas, which underwent a rearrangement of their transcriptome as extensive as in the striatum. Although tissue-enriched genes were downregulated in both areas, a neuroinflammation signature was only clearly induced in the R6/1 retina in which the observed glial activation was reminiscent of the situation in HD patient's brains. The retinal neuroinflammation was confirmed in the slow progressive knock-in zQ175 strain. Overall, these results demonstrated the suitability of the mouse retina as a research model for HD and its associated glial activation.

Changes in arterial myocyte excitability induced by subarachnoid hemorrhage in a rat model.

Aneurismal subarachnoid hemorrhage (aSAH) is a neurovascular disease produced by the rupture of the cerebral arteries and the extravasation of blood to the subarachnoid space and is accompanied by severe comorbidities. Secondarily associated vasospasm is one of the main side effects after hydrocephalus and possible rebleeding. Here, we analyze the alterations in function in the arteries of a rat model of SAH. For this, autologous blood was injected into the cisterna magna. We performed electrophysiological, microfluorimetric, and molecular biology experiments at different times after SAH to determine the functional and molecular changes induced by the hemorrhage. Our results confirmed that in SAH animals, arterial myocytes were depolarized on days 5 and 7, had higher [Ca2+]i on baseline, peaks and plateaus, and were more excitable at low levels of depolarization on day 7, than in the control and sham animals. Microarray analysis showed that, on day 7, the sets of genes related to voltage-dependent Ca2+ channels and K+ dynamics in SAH animals decreased, while the voltage-independent Ca2+ dynamics genes were over-represented. In conclusion, after SAH, several mechanisms involved in arterial reactivity were altered in our animal model, suggesting that there is no unique cause of vasospasm and alterations in several signaling pathways are involved in its development.

Changes in Adhesion and the Expression of Adhesion Molecules in PBMCs after Aneurysmal Subarachnoid Hemorrhage: Relation to Cerebral Vasospasm.

Aneurysmal subarachnoid hemorrhage (aSAH) is a neurovascular disease produced by extravasation of blood to the subarachnoid space after rupture of the cerebral vessels. After bleeding, the immune response is activated. The role of peripheral blood mononuclear cells (PBMCs) in this response is a current subject of research. We have analysed the changes in PBMCs of patients with aSAH and their interaction with the endothelium, focusing on their adhesion and the expression of adhesion molecules. Using an in vitro adhesion assay, we observed that the adhesion of PBMCs of patients with aSAH is increased. Flow cytometry analysis shows that monocytes increased significantly in patients, especially in those who developed vasospasm (VSP). In aSAH patients, the expression of CD162, CD49d, CD62L and CD11a in T lymphocytes and of CD62L in monocytes increased. However, the expression of CD162, CD43, and CD11a decreased in monocytes. Furthermore, monocytes from patients who developed arteriographic VSP had lower expression of CD62L. In conclusion, our results confirm that after aSAH, monocyte count and adhesion of PBMCs increase, especially in patients with VSP, and that the expression of several adhesion molecules is altered. These observations can help predict VSP and to improve the treatment of this pathology.

Unravelling the Inflammatory Processes in the Early Stages of Diabetic Nephropathy and the Potential Effect of (Ss)-DS-ONJ.

Inflammatory processes play a central role in the pathogenesis of diabetic nephropathy (DN) in the early stages of the disease. The authors demonstrate that the glycolipid mimetic (Ss)-DS-ONJ is able to abolish inflammation via the induction of autophagy flux and provokes the inhibition of inflammasome complex in ex vivo and in vitro models, using adult kidney explants from BB rats. The contribution of (Ss)-DS-ONJ to reducing inflammatory events is mediated by the inhibition of classical stress kinase pathways and the blocking of inflammasome complex activation. The (Ss)-DS-ONJ treatment is able to inhibit the epithelial-to-mesenchymal transition (EMT) progression, but only when the IL18 levels are reduced by the treatment. These findings suggest that (Ss)-DS-ONJ could be a novel, and multifactorial treatment for DN.

A New Perspective on Huntington's Disease: How a Neurological Disorder Influences the Peripheral Tissues.

Huntington's disease (HD) is a neurodegenerative disorder caused by a toxic, aggregation-prone expansion of CAG repeats in the HTT gene with an age-dependent progression that leads to behavioral, cognitive and motor symptoms. Principally affecting the frontal cortex and the striatum, mHTT disrupts many cellular functions. In fact, increasing evidence shows that peripheral tissues are affected by neurodegenerative diseases. It establishes an active crosstalk between peripheral tissues and the brain in different neurodegenerative diseases. This review focuses on the current knowledge of peripheral tissue effects in HD animal and cell experimental models and identifies biomarkers and mechanisms involved or affected in the progression of the disease as new therapeutic or early diagnostic options. The particular changes in serum/plasma, blood cells such as lymphocytes, immune blood cells, the pancreas, the heart, the retina, the liver, the kidney and pericytes as a part of the blood-brain barrier are described. It is important to note that several changes in different mouse models of HD present differences between them and between the different ages analyzed. The understanding of the impact of peripheral organ inflammation in HD may open new avenues for the development of novel therapeutic targets.

Neuropeptide Y facilitates P2X1 receptor-dependent vasoconstriction via Y1 receptor activation in small mesenteric arteries during sympathetic neurogenic responses.

ATP, norepinephrine and NPY are co-released by sympathetic nerves innervating arteries. ATP elicits vasoconstriction via activation of smooth muscle P2X receptors. The functional interaction between neuropeptide Y (NPY) and P2X receptors in arteries is not known. In this study we investigate the effect of NPY on P2X1-dependent vasoconstriction in mouse mesenteric arteries. Suramin or P2X1 antagonist NF449 abolished α,ß-meATP evoked vasoconstrictions. NPY lacked any direct vasoconstrictor effect but facilitated the vasoconstrictive response to α,ß-meATP. Mesenteric arteries expressed Y1 and Y4 receptors, but not Y2 or Y5. Y1 receptor inhibition (BIBO3304) reversed NPY facilitation of the α,ß-meATP-evoked vasoconstriction. L-type Ca2+ channel antagonism (nifedipine) had no effect on α,ß-meATP-evoked vasoconstrictions, but completely reversed NPY facilitation. Electrical field stimulation evoked sympathetic neurogenic vasoconstriction. Neurogenic responses were dependent upon dual α1-adrenergic (prazosin) and P2X1 (NF449) receptor activation. Y1 receptor antagonism partially reduced neurogenic vasoconstriction. Isolation of the P2X1 component by α1-adrenergic blockade allowed faciliatory effects of Y1 receptor activation to be explored. Y1 receptor antagonism reduced the P2X1 receptor component during neurogenic vasoconstriction. α1-adrenergic and P2X1 receptors are post-junctional receptors during sympathetic neurogenic vasoconstriction in mesenteric arteries. In conclusion, we have identified that NPY lacks a direct vasoconstrictor effect in mesenteric arteries but can facilitate vasoconstriction by enhancing the activity of P2X1, following activation by exogenous agonists or during sympathetic nerve stimulation. The mechanism of P2X1 facilitation by NPY involved activation of the NPY Y1 receptor and the L-type Ca2+ channel.

PKCα-Mediated Downregulation of RhoA Activity in Depolarized Vascular Smooth Muscle: Synergistic Vasorelaxant Effect of PKCα and ROCK Inhibition.

BACKGROUND/AIMS: Protein kinase C (PKC)- and RhoA/Rho-associated kinase (ROCK) play important roles in arterial sustained contraction. Although depolarization-elicited RhoA/ROCK activation is accepted, the role of PKC in depolarized vascular smooth muscle cells (VSMCs) is a subject of controversy. Our aim was to study the role of PKC in arterial contraction and its interaction with RhoA/ROCK. METHODS: Mass spectrometry was used to identify the PKC isoenzymes. PKCα levels and RhoA activity were analyzed by western blot and G-LISA, respectively, and isometric force was measured in arterial rings. RESULTS: In depolarized VSMCs RhoA and PKCα were translocated to the plasma membrane, where they colocalize and coimmunoprecipitate. Interestingly, depolarization-induced RhoA activation was downregulated by PKCα, effect reverted by PKCα inhibition. Phorbol 12,13-dibutyrate (PDBu) induced the translocation of PKCα to the plasma membrane, increased the level of RhoA in the cytosol and reduced RhoA/ROCK activity. These effects were reverted when PKC was inhibited. Pharmacological or siRNA inhibition of PKCα synergistically potentiated the vasorelaxant effect of RhoA/ROCK inhibition. CONCLUSION: The present study provides the first evidence that RhoA activity is downregulated by PKCα in depolarized and PDBu treated freshly isolated VSMCs and arteries, with an important physiological role on arterial contractility.

Relation of RhoA in Peripheral Blood Mononuclear Cells With Severity of Aneurysmal Subarachnoid Hemorrhage and Vasospasm.

BACKGROUND AND PURPOSE: Rho-kinase, an effector of RhoA, is associated with various cardiovascular diseases in circulating blood cells. However, the role of RhoA/Rho-kinase in peripheral blood mononuclear cells from patients with spontaneous aneurysmal subarachnoid hemorrhage (aSAH) has not yet been studied in relation to the severity of this disease. Therefore, we analyzed the expression and activity of RhoA as a possible biomarker in aSAH. METHODS: Twenty-four patients with aSAH and 15 healthy subjects were examined. Peripheral blood mononuclear cells were collected, and RhoA activity and expression were determined by RhoA activation assay kit (G-LISA) and enzyme-linked immunosorbent assay tests, respectively. The severity of aSAH was determined from the World Federation of Neurological Surgeon scale, and vasospasm was evaluated using clinical symptoms, arteriography, and sonography. RESULTS: RhoA expression was significantly increased in peripheral blood mononuclear cells from patients on days 0, 2, and 4 after aSAH versus healthy subjects (P=0.036, 0.010, and 0.018, respectively, by U Mann-Whitney analysis). There was a significant correlation between RhoA expression and injury severity on days 2 and 4 (Spearman test, day 2: r=0.682, n=14, P=0.007; day 4: r=0.721, n=14, P=0.004). No significant correlation was observed on day 0 (day 0: r=0.131, n=6, P=0.805). Active RhoA was not significantly different in patients and healthy subjects on days 0, 2, and 4 (P=0.243, 0.222, and 0.600, respectively) nor did it increase significantly on days 0 and 2 in patients with vasospasm versus patients without vasospasm (P=0.064 and 0.519, respectively). In contrast, active RhoA was significantly higher on day 4 in patients who developed vasospasm versus patients without vasospasm (P=0.028). CONCLUSIONS: Our preliminary results indicate that RhoA expression and activity in peripheral blood mononuclear cells might be related with aSAH severity and cerebral vasospasm. RhoA is a potential biomarker of the risks associated with aSAH.

Role of L-type Ca(2+) channels, sarcoplasmic reticulum and Rho kinase in rat basilar artery contractile properties in a new model of subarachnoid hemorrhage.

We have previously described that L-type Ca(2+) channels' (LTCCs) activation and metabotropic Ca(2+) release from the sarcoplasmic reticulum (SR) regulate RhoA/Rho kinase (ROCK) activity and sustained arterial contraction. We have investigated whether this signaling pathway can be altered in a new experimental model of subarachnoid hemorrhage (SAH). For this purpose, arterial reactivity was evaluated on days 1 to 5 after surgery. A significant increase of basal tone, measured 4 and 60min after normalization, was observed on day 5 after SAH and at 60min on days 2 and 3 after SAH. This phenomenon was suppressed with LTCCs and ROCK inhibitors. We have also studied arterial rings vasoreactivity in response to high K(+) solutions. Interestingly, there were no significant differences in the phasic component of the high K(+)-induced contraction between sham and SAH groups, whereas a significant increase in the sustained contraction was observed on day 5 after SAH. This latter component was sensitive to fasudil, and selectively reduced by low nifedipine concentration, and phospholipase C and SR-ATPase inhibitors. Therefore, our data suggest that the metabotropic function of LTCCs is potentiated in SAH. Our results could provide a new strategy to optimize the pharmacological treatment of this pathological process.

Contractile responses to rat urotensin II in resting and depolarized basilar arteries

The effects of human urotensin II (hUII) on the vascular tone of different animal species has been studied extensively. However, little has been reported on the vasoactive effects of rat urotensin (rUII) in murine models. The aim of the present study was to investigate the effects of rUII on vasoreactivity in rat basilar arteries. Basilar arteries from adult male Wistar rats (300-350 g) were isolated, cut in rings, and mounted on a small vessel myograph to measure isometric tension. rUII concentrations were studied in both resting and depolarized state. To remove endothelial nitric oxide effects from the rUII response, we treated selected arterial rings with Nω-nitro-L-arginine methyl ester (L-NAME). 10 μM rUII produced a potent vasoconstrictor response in rat basilar arteries with intact endothelium, while isometric forces remained unaffected in arterial rings treated with lower rUII concentrations. Although L-NAME did not have a significant effect on 10 μM rUII-evoked contraction, it slightly increased arterial ring contraction elicited by 1 μM rUII. In depolarized arteries, dose-dependent rUII increased depolarization-induced contractions. This effect was suppressed by L-NAME. Our results show that the rat basilar artery has a vasoconstrictor response to rUII. The most potent vasoconstrictor effect was produced by lower doses of rUII (0.1 and 1 μM) in depolarized arteries with intact endothelium. This effect could facilitate arterial vasospasm in vascular pathophysiological processes such as subarachnoid hemorrhage and hypertension, when sustained depolarization and L-type Ca2+ channel activation are present

Contractile responses to rat urotensin II in resting and depolarized basilar arteries.

The effects of human urotensin II (hUII) on the vascular tone of different animal species has been studied extensively. However, little has been reported on the vasoactive effects of rat urotensin (rUII) in murine models. The aim of the present study was to investigate the effects of rUII on vasoreactivity in rat basilar arteries. Basilar arteries from adult male Wistar rats (300-350 g) were isolated, cut in rings, and mounted on a small vessel myograph to measure isometric tension. rUII concentrations were studied in both resting and depolarized state. To remove endothelial nitric oxide effects from the rUII response, we treated selected arterial rings with Nω-nitro-L-arginine methyl ester (L-NAME). 10 µM rUII produced a potent vasoconstrictor response in rat basilar arteries with intact endothelium, while isometric forces remained unaffected in arterial rings treated with lower rUII concentrations. Although L-NAME did not have a significant effect on 10 µM rUII-evoked contraction, it slightly increased arterial ring contraction elicited by 1 µM rUII. In depolarized arteries, dose-dependent rUII increased depolarization-induced contractions. This effect was suppressed by L-NAME. Our results show that the rat basilar artery has a vasoconstrictor response to rUII. The most potent vasoconstrictor effect was produced by lower doses of rUII (0.1 and 1 µM) in depolarized arteries with intact endothelium. This effect could facilitate arterial vasospasm in vascular pathophysiological processes such as subarachnoid hemorrhage and hypertension, when sustained depolarization and L-type Ca(2+) channel activation are present.