Long term remodeling of rat pial microcirculation after transient middle cerebral artery occlusion and reperfusion.

Objective: The aim of this study was to assess the in vivo structural and functional remodeling of pial arteriolar networks in the ischemic area of rats submitted to transient middle cerebral artery occlusion (MCAO) and different time intervals of reperfusion. Methods and results: Two closed cranial windows were implanted above the left and right parietal cortex to observe pial microcirculation by fluorescence microscopy. The geometric characteristics of pial arteriolar networks, permeability increase, leukocyte adhesion and capillary density were analyzed after 1 h or 1, 7, 14 or 28 days of reperfusion. MCAO and 1-hour reperfusion caused marked microvascular changes in pial networks. The necrotic core was devoid of vessels, while the penumbra area presented a few arterioles, capillaries and venules with severe neuronal damage. Penumbra microvascular permeability and leukocyte adhesion were pronounced. At 7 days of reperfusion, new pial arterioles were organized in anastomotic vessels, overlapping the ischemic core and in penetrating pial arterioles. Vascular remodeling caused different arteriolar rearrangement up to 28 days of reperfusion and animals gradually regained their motor and sensory functions. Conclusions: Transient MCAO-induced pial-network remodeling is characterized by arteriolar anastomotic arcades. Remodeling mechanisms appear to be accompanied by an increased expression of nitric oxide synthases

Effects of bone marrow mesenchymal stem cells (BM-MSCs) on rat pial microvascular remodeling after transient middle cerebral artery occlusion

Previous studies have shown that the pial microcirculation remodeling improves neurological outcome after middle cerebral artery occlusion (MCAO), accompanied by higher expression of vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS), modulating in vivo angiogenesis. This study was aimed to assess the effects of bone marrow mesenchymal stem cells (BM-MSCs) infused after MCAO on rat pial microcirculation. Animals were subjected to 2 h MCAO followed by BM-MSCs infusion into internal carotid artery. Pial microcirculation was observed at different reperfusion times by fluorescence microscopy. Geometric characteristics of arteriolar networks, permeability increase, leukocyte adhesion, perfused capillary density, VEGF, and endothelial nitric oxide synthase (e-NOS) expression were evaluated. Green fluorescent protein (GFP)-BM-MSCs were used to evaluate their distribution and cell phenotype development during reperfusion. BM-MSCs stimulated a geometric rearrangement of pial networks with formation of new anastomotic vessels sprouting from preexistent arterioles in the penumbra at 7-14-28 days of reperfusion. At the same time VEGF and eNOS expression increased. GFP-BM-MSCs appear to be involved in endothelial and smooth muscle cell programming in the infarcted area. In conclusion, transient MCAO induced pial vascular remodeling characterized by arteriolar anastomotic arcades (originated from preexistent arterioles in penumbra area) able to overlap the ischemic core supplying blood to the neuronal tissue. BM-MSCs appear to accelerate angiogenic processes facilitating new vessel formation; this mechanism was promoted by an increase in VEGF and eNOS expression

Myricetin preserves rat pial microcirculation from injury induced by cerebral hypoperfusion and reperfusion

Background/Objective: Myricetin, a flavonoid compound, is widely diffused in vegetables, fruits and beverages, well known for its antioxidant and anti-inflammatory properties. The present study was aimed to investigate the acute effects of myricetin on the pial microvascular alterations and oxygen-derived free radical formation, due to 30 min cerebral blood flow lowering (CBFL) and subsequent cerebral blood flow resumption (CBFR). Methods: Rat pial microvasculature was investigated using fluorescence microscopy through a closed cranial window. At first, arterioles were classified according to the Strahler’s ordering scheme. Then, arteriolar diameter, permeability increase, leukocyte adhesion to venular walls, perfused capillary length (CPL) and red blood cell velocity (VRBC) were quantified by computerized methods. Finally, reactive oxygen species (ROS) production was investigated in vivo by 2′-7′-dichlorofluoresceindiacetate assay and infarct size by 2,3,5-triphenyltetrazolium chloride staining. Results: After 30 min CBFL and 60 min CBFR, a decrease of arteriolar diameter, CPL and VRBC was detected; furthermore, increases in microvascular leakage and leukocyte adhesion were observed in hypoperfused animals. Conversely, myricetin administration induced dose-related arteriolar dilation, reduction in microvascular permeability as well as leukocyte adhesion when compared to those detected in bilateral common carotid artery occlusion-submitted animals; moreover, CPL and VRBC were preserved. In animals treated with myricetin the ROS production was blunted and infarct size significantly reduced. Conclusion: In conclusion, myricetin acute administration showed dose-related protective effects on rat pial microcirculation during CBFL and subsequent CBFR, inducing arteriolar dilation and inhibiting ROS formation, consequently preserving the blood brain barrier integrity

Arterial Network Geometric Characteristics and Regulation of Capillary Blood Flow in Hamster Skeletal Muscle Microcirculation

This study was aimed to characterize the geometric arrangement of hamster skeletal muscle arteriolar networks and to assess the in vivo rhythmic diameter changes of arterioles to clarify regulatory mechanisms of the capillary perfusion. The experimental study was carried out in male Syrian hamsters implanted with a plastic chamber in the dorsum skin under pentobarbital anesthesia. The skeletal muscle microvessels were visualized by fluorescence microscopy. The vessel diameters, lengths and the rhythmic diameter changes of arterioles were analyzed with computer-assisted techniques. The arterioles were classified according to a centripetal ordering scheme. In hamster skeletal muscle microvasculature the terminal branchings, differentiated in long and short terminal arteriolar trees (TATs), originated from anastomotic vessels, defined “arcading” arterioles. The long TATs presented different frequencies along the branching vessels; order 4 arterioles had frequencies lower than those observed in the order 3, 2, and 1 vessels. The short TAT order 3 arterioles, directly originating from “arcading” parent vessels, showed a frequency dominating all daughter arterioles. The amplitude of diameter variations in larger vessels was in the range 30–40% of mean diameter, while it was 80–100% in order 3, 2, and 1 vessels. Therefore, the complete constriction of arterioles, caused an intermittent capillary blood perfusion. L-arginine or papaverine infusion caused dilation of arterioles and transient disappearing of vasomotion waves and induced perfusion of all capillaries spreading from short and long TAT arrangements. Therefore, the capillary blood flow was modulated by changes in diameter of terminal arterioles penetrating within the skeletal muscle fibers, facilitating redistribution of blood flow according to the metabolic demands of tissues

Malvidin's Effects on Rat Pial Microvascular Permeability Changes Due to Hypoperfusion and Reperfusion Injury

The present study was aimed to evaluate the malvidin's protective effects on damage induced by 30 min bilateral common carotid artery occlusion (BCCAO) and 60 min reperfusion (RE) in rat pial microcirculation. Rat pial microcirculation was observed using fluorescence microscopy through a closed cranial window. Western blotting analysis was performed to investigate the endothelial nitric oxide synthase (eNOS), phosphorylated eNOS (p-eNOS) and matrix metalloproteinase 9 (MMP-9) expression. Moreover, MMP-9 activity was evaluated by zymography. Finally, neuronal damage and radical oxygen species (ROS) formation were assessed. In all animals, pial arterioles were classified in five orders of branching according to Strahler's method. In hypoperfused rats, 30 min BCCAO and 60 min RE caused a decrease in arteriolar diameter, an increase in microvascular leakage and leukocyte adhesion, accompanied by decreased capillary perfusion and red blood cell velocity (VRBC). Moreover, marked neuronal damage and evident ROS generation were detected. Conversely, malvidin administration induced arteriolar dilation in dose-related manner, reducing microvascular leakage as well as leukocyte adhesion. Capillary perfusion and VRBC were protected. Nitric oxide (NO) synthase inhibition significantly attenuated malvidin's effects on arteriolar diameter. Western blotting analysis revealed an increase in eNOS and p-eNOS expression, while zymography indicated a decrease in MMP-9 activity after malvidin's administration. Furthermore, malvidin was able to prevent neuronal damage and to decrease ROS generation. In conclusion, malvidin protects rat pial microcirculation against BCCAO/RE injury, preventing blood-brain impairment and neuronal loss. Malvidin's effects appear to be mediated by eNOS activation and scavenger activity

Microvascular blood flow improvement in hyperglycemic obese adult patients by hypocaloric diet

The present study was aimed to assess the changes in skin microvascular blood flow (SBF) in newly diagnosed hyperglycemic obese subjects, administered with hypocaloric diet. Adult patients were recruited and divided in three groups: NW group (n=54), NG (n=54) and HG (n=54) groups were constituted by normal weight, normoglycemic and hyperglycemic obese subjects, respectively. SBF was measured by laser Doppler perfusion monitoring technique and oscillations in blood flow were analyzed by spectral methods under baseline conditions, at 3 and 6 months of dietary treatment. Under resting conditions, SBF was lower in HG group than in NG and NW ones. Moreover, all subjects showed blood flow oscillations with several frequency components. In particular, hyperglycemic obese patients revealed lower spectral density in myogenic-related component than normoglycemic obese and normal weight ones. Moreover, postocclusive reactive hyperemia (PORH) was impaired in hyperglycemic obese compared to normoglycemic and normal weigh subjects. After hypocaloric diet, in hyperglycemic obese patients there was an improvement in SBF accompanied by recovery in myogenic-related oscillations and arteriolar responses during PORH. In conclusion, hyperglycemia markedly affected peripheral microvascular function; hypocaloric diet ameliorated tissue blood flow

Oscillazioni del flusso microvascolare in pazienti obesi iperglicemici

Aim: The present study was aimed to investigate skin microvascular blood flow oscillations by noninvasively LDPM and to evaluate the effects of insulin resistance and hyperglycemia on skin perfusion in obese subjects. Furthermore, we assessed the effects of hypocaloric diet on skin microvascular blood flow in newly diagnosed hyperglicemic obese patients. Methods: Fifty-four normoglycemic (27 females) and fifty-four hyperglycemic (27 females) obese adults were recruited from our Outpatient Clinic and treated with hypocaloric diet for 6 months. Skin microvascular blood flow was recorded by laser Doppler flowmeter and blood flow oscillations were analyzed by wavelet transform. Moreover, reactive hyperemia, due to 2 min brachial artery occlusion, was investigated in fourteen normoglycemic (7 females) and sixteen hyperglycemic (8 females) obese adults. Results: Skin microvascular blood flow (9.4 ± 0.3 vs 11.8 ± 0.4 PU, p<0.01) and total power spectral density (PSD: 113.2 ± 10.1 vs 176.5 ± 11.4 PU2/Hz, p<0.01) were lower in the hyperglycemic than normoglycemic people. Among the frequency components of blood flow oscillations, myogenic activity spectral density was lower in hyperglycemic than normoglycemic patients (31.0 ± 0.7 vs 34.9 ± 0.7 %, p<0.01). Moreover, hyperglycemic subjects showed lower hyperemic response to artery occlusion compared to the corresponding one observed in normoglycemic patients. After 6 month hypocaloric diet, blood flow and spectral denisty significantly increased in hyperglycemic subjects. Myogenic activity-related oscillatory component spectral density was enhanced, while post-occlusive reactive hyperemia significantly improved. Conclusions: In conclusion, insulin resistance- and hyperglycemia-associated microvascular alterations severely affect peripheral resistance and tissue perfusion. Hypocaloric diet appeared to ameliorate arteriolar dilation and blood flow redistribution to tissue

Pial microvascular responses induced by transient bilateral common carotid artery occlusion in Zucker rats

This study was aimed to assess the in vivo geometric and functional characteristics of lean Zucker (ZL) and obese Zucker rat (ZO) pial microvascular networks and to evaluate the vascular responses to cerebral hypoperfusion-reperfusion. Rat pial microcirculation was observed by fluorescence microscopy through a closed cranial window. Bilateral common carotid artery occlusion (BCCAO) lasted 30 min and reperfusion 60 min. Arterioles were classified according to Strahler's ordering scheme. Arteriolar diameter was determined by computer assisted-method as well as permeability increase, leukocyte adhesion and perfused capillary length. Neuronal damage was evaluated by TTC staining. ZO rats did not show order 5 vessels; ZO pial arterioles showed high asymmetry in the largest vessels and reduced number of branchings compared with those detected in ZL and Wistar rats. BCCAO and reperfusion caused more severe microvascular damages in ZO compared with ZL and Wistar rats. Vascular responses to acetylcholine and papaverine in ZO rats were significantly reduced compared with Wistar and ZL rats under baseline condition and at the end of reperfusion. Moreover, ZO rats showed more pronounced lesion in the cortex and striatum. Obesity and hyperglycemia could increase vascular remodeling in cerebral networks, with elevated risk of adverse outcome after brain hypoperfusion-reperfusion

Pial microvascular responses induced by transient bilateral common carotid artery occlusion in zucker rats.

This study was aimed to assess the in vivo geometric and functional characteristics of lean Zucker (ZL) and obese Zucker rat (ZO) pial microvascular networks and to evaluate the vascular responses to cerebral hypoperfusion-reperfusion. Rat pial microcirculation was observed by fluorescence microscopy through a closed cranial window. Bilateral common carotid artery occlusion (BCCAO) lasted 30 min and reperfusion 60 min. Arterioles were classified according to Strahler's ordering scheme. Arteriolar diameter was determined by computer assisted-method as well as permeability increase, leukocyte adhesion and perfused capillary length. Neuronal damage was evaluated by TTC staining. ZO rats did not show order 5 vessels; ZO pial arterioles showed high asymmetry in the largest vessels and reduced number of branchings compared with those detected in ZL and Wistar rats. BCCAO and reperfusion caused more severe microvascular damages in ZO compared with ZL and Wistar rats. Vascular responses to acetylcholine and papaverine in ZO rats were significantly reduced compared with Wistar and ZL rats under baseline condition and at the end of reperfusion. Moreover, ZO rats showed more pronounced lesion in the cortex and striatum. Obesity and hyperglycemia could increase vascular remodeling in cerebral networks, with elevated risk of adverse outcome after brain hypoperfusion-reperfusion

Microvascular responses to aldosterone in hamster cheek pouch microcirculation

The aim of the present study was to assess the in vivo effects of aldosterone topically applied on the hamster cheek pouch microcirculation under baseline conditions or during ischemia-reperfusion. Male Syrian hamsters were anesthetized, tracheotomized and intubated. They were studied under baseline conditions or submitted to ischemia-reperfusion. Cheek pouch microvessels were visualized by fluorescence microscopy. Microvascular parameters were determined by computerized methods. Under baseline conditions, aldosterone (0.2, 0.5, 2.4 μM/L/2 min) induced dose-dependent constriction of all arterioles within 2.0 ± 0.5 min of administration. Diameter reduction was in the same range in smaller arterioles: A3 ones constricted by 24 ± 3% of baseline (at the highest dose). Aldosterone applied prior to ischemia and at reperfusion caused arteriolar constriction, marked microvascular permeability (0.66 ± 0.03 Normalized Grey Level), reduction in perfused capillary (-70 ± 4% of baseline) and leukocyte adhesion. All changes were statistically significant compared with ischemic animals. Potassium canrenoate (mineralcorticoid receptor inhibitor) prior to aldosterone did not abolish the aldosterone-induced effects, while valsartan (angiotensin II AT1 receptor inhibitor) prior to aldosterone ameliorated microvascular ischemia-reperfusion injury. In conclusion, aldosterone determined dose-dependent arteriolar constriction likely by angiotensin II type-1 receptor activation (non-genomic mechanism) worsening the effects of ischemia-reperfusion on capillary perfusion, while protecting from free radical formation