Beneficial effects of empagliflozin and liraglutide on the cerebral microcirculation of diabetic rats.

OBJECTIVES: This study aimed to evaluate the effects of the antidiabetics liraglutide, a GLP-1 analog, and empagliflozin, an SGLT-2 inhibitor, on the brain microcirculation of diabetic rats. METHODS: Type 2 diabetes mellitus (DM) was experimentally induced in male Wistar rats by combining a high-fat diet and a low dose of streptozotocin (35 mg/kg). Liraglutide (100 µg/kg s.c.) and empagliflozin (10 mg/kg, oral) were administered for 5 weeks. Body weight was monitored periodically. Oral glucose tolerance, fasting glycemia, and blood triglycerides were evaluated after the treatments. Endothelial-leukocyte interactions in the brain microcirculation and structural capillary density were assessed. RESULTS: DM rats presented metabolic and cerebrovascular alterations. Liraglutide treatment decreased body weight and blood triglycerides of DM rats. Empagliflozin treatment improved glucose tolerance but only the combination therapy significantly reduced fasting blood glucose. Both treatments and their combination reduced leukocyte adhesion into the endothelium of brain venules. However, empagliflozin was more effective in preventing DM-induced microvascular rarefaction. CONCLUSION: These findings suggest that chronic treatment with SGLT2 inhibitors and GLP-1 receptor agonists may serve as potential therapeutic approaches to prevent microvascular complications associated with diabetes.

TLR4 mutation protects neurovascular function and cognitive decline in high-fat diet-fed mice.

BACKGROUND: Metabolic syndrome (MS) is defined as a low-grade proinflammatory state in which abnormal metabolic and cardiovascular factors increase the risk of developing cardiovascular disease and neuroinflammation. Events, such as the accumulation of visceral adipose tissue, increased plasma concentrations of free fatty acids, tissue hypoxia, and sympathetic hyperactivity in MS may contribute to the direct or indirect activation of Toll-like receptors (TLRs), specifically TLR4, which is thought to be a major component of this syndrome. Activation of the innate immune response via TLR4 may contribute to this state of chronic inflammation and may be related to the neuroinflammation and neurodegeneration observed in MS. In this study, we investigated the role of TLR4 in the brain microcirculation and in the cognitive performance of high-fat diet (HFD)-induced MS mice. METHODS: Wild-type (C3H/He) and TLR4 mutant (C3H/HeJ) mice were maintained under a normal diet (ND) or a HFD for 24 weeks. Intravital video-microscopy was used to investigate the functional capillary density, endothelial function, and endothelial-leukocyte interactions in the brain microcirculation. Plasma concentrations of monocyte chemoattractant protein-1 (MCP-1), adipokines and metabolic hormones were measured with a multiplex immunoassay. Brain postsynaptic density protein-95 and synaptophysin were evaluated by western blotting; astrocytic coverage of the vessels, microglial activation and structural capillary density were evaluated by immunohistochemistry. RESULTS: The HFD-induced MS model leads to metabolic, hemodynamic, and microcirculatory alterations, as evidenced by capillary rarefaction, increased rolling and leukocyte adhesion in postcapillary venules, endothelial dysfunction, and less coverage of astrocytes in the vessels, which are directly related to cognitive decline and neuroinflammation. The same model of MS reproduced in mice deficient for TLR4 because of a genetic mutation does not generate such changes. Furthermore, the comparison of wild-type mice fed a HFD and a normolipid diet revealed differences in inflammation in the cerebral microcirculation, possibly related to lower TLR4 activation. CONCLUSIONS: Our results demonstrate that TLR4 is involved in the microvascular dysfunction and neuroinflammation associated with HFD-induced MS and possibly has a causal role in the development of cognitive decline.

Systemic microvascular endothelial dysfunction and disease severity in COVID-19 patients: Evaluation by laser Doppler perfusion monitoring and cytokine/chemokine analysis.

BACKGROUND: Microvascular dysfunction, serum cytokines and chemokines may play important roles in pathophysiology of coronavirus disease 2019 (COVID-19), especially in severe cases. METHODS: Patients with COVID-19 underwent non-invasive evaluation of systemic endothelium-dependent microvascular reactivity - using laser Doppler perfusion monitoring in the skin of the forearm - coupled to local thermal hyperemia. Maximal microvascular vasodilatation (44 °C thermal plateau phase) was used as endpoint. A multiplex biometric immunoassay was used to assess a panel of 48 serum cytokines and chemokines. Severe COVID-19 (S-COVID) was defined according to WHO criteria, while all other cases of COVID-19 were considered mild to moderate (M-COVID). A group of healthy individuals who tested negative for SARS-CoV-2 served as a control group and was also evaluated with LDPM. RESULTS: Thirty-two patients with COVID-19 (25% S-COVID) and 14 controls were included. Basal microvascular flow was similar between M-COVID and controls (P = 0.69) but was higher in S-COVID than in controls (P = 0.005) and M-COVID patients (P = 0.01). The peak microvascular vasodilator response was markedly decreased in both patient groups (M-COVID, P = 0.001; S-COVID, P < 0.0001) compared to the healthy group. The percent increases in microvascular flow were markedly reduced in both patient groups (M-COVID, P < 0.0001; S-COVID, P < 0.0001) compared to controls. Patients with S-COVID had markedly higher concentrations of dissimilar proinflammatory cytokines and chemokines, compared to patients with M-COVID. CONCLUSIONS: In patients with COVID-19, especially with S-COVID, endothelium-dependent microvascular vasodilator responses are reduced, while serum cytokines and chemokines involved in the regulation of vascular function and inflammation are increased.

Physical exercise promotes astrocyte coverage of microvessels in a model of chronic cerebral hypoperfusion.

BACKGROUND: Brain circulation disorders such as chronic cerebral hypoperfusion have been associated with a decline in cognitive function during the development of dementia. Astrocytes together with microglia participate in the immune response in the CNS and make them potential sentinels in the brain parenchyma. In addition, astrocytes coverage integrity has been related to brain homeostasis. Currently, physical exercise has been proposed as an effective intervention to promote brain function improvement. However, the neuroprotective effects of early physical exercise on the astrocyte communication with the microcirculation and the microglial activation in a chronic cerebral hypoperfusion model are still unclear. The aim of this study was to investigate the impact of early intervention with physical exercise on cognition, brain microcirculatory, and inflammatory parameters in an experimental model of chronic cerebral hypoperfusion induced by permanent bilateral occlusion of the common carotid arteries (2VO). METHODS: Wistar rats aged 12 weeks were randomly divided into four groups: Sham-sedentary group (Sham-Sed), Sham-exercised group (Sham-Ex), 2VO-sedentary group (2VO-Sed), and 2VO-exercised group (2VO-Ex). The early intervention with physical exercise started 3 days after 2VO or Sham surgery during 12 weeks. Then, the brain functional capillary density and endothelial-leukocyte interactions were evaluated by intravital microscopy; cognitive function was evaluated by open-field test; hippocampus postsynaptic density protein 95 and synaptophysin were evaluated by western blotting; astrocytic coverage of the capillaries, microglial activation, and structural capillary density were evaluated by immunohistochemistry. RESULTS: Early moderate physical exercise was able to normalize functional capillary density and reduce leukocyte rolling in the brain of animals with chronic cerebral hypoperfusion. These effects were accompanied by restore synaptic protein and the improvement of cognitive function. In addition, early moderate exercise improves astrocytes coverage in blood vessels of the cerebral cortex and hippocampus, decreases microglial activation in the hippocampus, and improves structural capillaries in the hippocampus. CONCLUSIONS: Microcirculatory and inflammatory changes in the brain appear to be involved in triggering a cognitive decline in animals with chronic cerebral ischemia. Therefore, early intervention with physical exercise may represent a preventive approach to neurodegeneration caused by chronic cerebral hypoperfusion.

Microvascular Effects of Echinodorus grandiflorus on Cardiovascular Disorders.

Echinodorus grandiflorus is a semiaquatic plant native to Brazil and belongs to the Alismataceae family. Infusion preparations of the leaves of this plant are often used due to its diuretic, blood pressure lowering, and anti-inflammatory properties. Our aim was to investigate the effects of chronic treatment with the crude hydroalcoholic extract of E. grandiflorus on central and peripheral microvascular changes induced in a model of hypertension and diabetes. The hemodynamic and microvascular effects of E. grandiflorus extract (50, 100, or 200 mg/kg/day for 28 days) or the isolated major diterpene from E. grandiflorus (3 to 10 mg/kg i. v.) were evaluated in spontaneously hypertensive rats using tail plethysmography and intravital fluorescence videomicroscopy, respectively, and were compared to vehicle-treated normotensive Wistar-Kyoto rats. We also investigated the protective effects of chronic treatment with E. grandiflorus (100 mg/kg/day) in brain capillary density and leukocyte-endothelium interactions on the brain vessels of DM-spontaneously (DM: diabetes mellitus) hypertensive rats. Chronically treating spontaneously hypertensive rats with increasing doses of crude hydroalcoholic E. grandiflorus extract resulted in significant dose-dependent reductions in systolic blood pressure and an anti-inflammatory effect on the brain microcirculation of DM-spontaneously hypertensive rat animals. Using laser speckle contrast imaging, we observed that intravenous administration of the major isolated clerodane diterpene metabolite (1 - 10 mg/kg) increased microvascular blood flow by 25% in spontaneously hypertensive rat skeletal muscle. The results of this study show that E. grandiflorus extracts can be useful in the prevention and reduction of microcirculatory damage in arterial hypertension and other diseases that involve microvascular dysfunction.

The Neurotropic Parasite Toxoplasma gondii Induces Sustained Neuroinflammation with Microvascular Dysfunction in Infected Mice.

Toxoplasmosis is one of the leading parasitic diseases worldwide. Some data suggest that chronic acquired toxoplasmosis could be linked to behavioral alterations in humans. The parasite infects neurons, forming immunologically silent cysts. Cerebral microcirculation homeostasis is determinant to brain functions, and pathologic states can alter capillarity or blood perfusion, leading to neurodegeneration and cognitive deficits. Albino mice were infected with Toxoplasma gondii (ME49 strain) and analyzed after 10, 40, and 180 days. Infected mice presented decreased cerebral blood flow at 10 and 40 days post infection (dpi), which were restored at 180 dpi, as shown by laser speckle contrast imaging. Intravital microscopy demonstrated that infection led to significant capillary rarefaction, accompanied by neuroinflammation, with microglial activation and increased numbers of rolling and adherent leukocytes to the wall of cerebral capillaries. Acetylcholine-induced vasodilation was altered at all time points, and blood brain barrier permeability was evident in infected animals at 40 dpi. Infection reduced angiogenesis, with a decreased number of isolectin B4-stained blood vessels and a decrease in length and branching of laminin-stained capillaries. Sulfadiazine reduced parasite load and partially repaired microvascular damages. We conclude that T. gondii latent infection causes a harmful insult in the brain, promoting neuroinflammation and microcirculatory dysfunction in the brain, with decreased angiogenesis and can contribute to a neurodegenerative process.

Acute simvastatin treatment restores cerebral functional capillary density and attenuates angiotensin II-induced microcirculatory changes in a model of primary hypertension.

OBJECTIVE: We investigated the acute effects of SIM on cerebral microvascular rarefaction and dysfunction in SHRs. METHODS: Male WKY and SHRs were divided into 4 groups of 8 animals each: WKY-CTL and SHR-CTL, treated with 0.9% saline; and WKY+SIM and SHR+SIM, treated with SIM (30 mg/kg/d) for 3 days by gavage. Cerebral FCD was assessed by intravital fluorescence videomicroscopy. mCBF before and after administration within the cranial window of angiotensin II (1 µmol L-1 ) was investigated using laser speckle contrast imaging. RESULTS: Cerebral FCD was reduced in SHR-CTL compared to WKY-CTL (P < .05). SIM increased cerebral FCD in SHRs compared to SHR-CTL (P < .05). The mCBF was reduced in SHR-CTL compared to WKY-CTL (P < .05), and SIM increased mCBF compared with SHR-CTL (P < .05). Angiotensin II elicited a reduction of mCBF in SHR-CTL and increased mCBF in WKY-CTL (SHR-CTL -13.53 ± 2% vs WKY-CTL +13.74 ± 4%; P < .001), which was attenuated in SHRs treated with SIM (SHR+SIM -6.7 ± 1% vs SHR-CTL -13.53 ± 2%; P < .01). CONCLUSIONS: The antihypertensive effect of SIM is associated with an improvement in cerebral microvascular perfusion and capillary density that may help to prevent hypertension-induced cerebrovascular damage independent of cholesterol-lowering.

Cerebral microvascular dysfunction in metabolic syndrome is exacerbated by ischemia-reperfusion injury.

BACKGROUND: Metabolic syndrome (MetS) is associated with an increased risk of cerebrovascular diseases, including cerebral ischemia. Microvascular dysfunction is an important feature underlying the pathophysiology of cerebrovascular diseases. In this study, we aimed to investigate the impacts of ischemia and reperfusion (IR) injury on the cerebral microvascular function of rats with high-fat diet-induced MetS. RESULTS: We examined Wistar rats fed a high-fat diet (HFD) or normal diet (CTL) for 20 weeks underwent 30 min of bilateral carotid artery occlusion followed by 1 h of reperfusion (IR) or sham surgery. Microvascular blood flow was evaluated on the parietal cortex surface through a cranial window by laser speckle contrast imaging, functional capillary density, endothelial function and endothelial-leukocyte interactions by intravital videomicroscopy. Lipid peroxidation was assessed by TBARs analysis, the expression of oxidative enzymes and inflammatory markers in the brain tissue was analyzed by real-time PCR. The cerebral IR in MetS animals induced a functional capillary rarefaction (HFD IR 117 ± 17 vs. CTL IR 224 ± 35 capillary/mm2; p < 0.05), blunted the endothelial response to acetylcholine (HFD IR -16.93% vs. CTL IR 16.19% from baseline inner diameter p < 0.05) and increased the endothelial-leukocyte interactions in the venules in the brain. The impact of ischemia on the cerebral microvascular blood flow was worsened in MetS animals, with a marked reduction of cerebral blood flow, exposing brain tissue to a higher state of hypoxia. CONCLUSIONS: Our results demonstrate that during ischemia and reperfusion, animals with MetS are more susceptible to alterations in the cerebral microcirculation involving endothelial dysfunction and oxidative stress events.

Statins prevent cognitive impairment after sepsis by reverting neuroinflammation, and microcirculatory/endothelial dysfunction.

Acute brain dysfunction is a frequent condition in sepsis patients and is associated with increased mortality and long-term neurocognitive consequences. Impaired memory and executive function are common findings in sepsis survivors. Although neuroinflammation and blood-brain barrier dysfunction have been associated with acute brain dysfunction and its consequences, no specific treatments are available that prevent cognitive impairment after sepsis. Experimental sepsis was induced in Swiss Webster mice by intraperitoneal injection of cecal material (5mg/kg, 500µL). Control groups (n=5/group each experiment) received 500µL of saline. Support therapy recover (saline 0.9%, 1mL and imipenem 30mg/kg) were applied (6, 24 and 48h post injection, n=5-10/group, each experiment), together or not with additive orally treatment with statins (atorvastatin/simvastatin 20mg/kg b.w.). Survival rate was monitored at 6, 24 and 48h. In a setting of experiments, animals were euthanized at 6 and 24h after induction for biochemical, immunohistochemistry and intravital analysis. Statins did not prevented mortality in septic mice, however survivors presented lower clinical score. At another setting of experiments, after 15days, mice survivors from fecal supernatant peritoneal sepsis presented cognitive dysfunction for contextual hippocampal and aversive amygdala-dependent memories, which was prevented by atorvastatin/simvastatin treatment. Systemic and brain tissue levels of proinflammatory cytokines/chemokines and activation of microglial were lower in septic mice treated with statins. Brain lipid peroxidation and myeloperoxidase levels were also reduced by statins treatment. Intravital examination of the brain vessels of septic animals revealed decreased functional capillary density and increased rolling and adhesion of leukocytes, and blood flow impairment, which were reversed by treatment with statins. In addition, treatment with statins restored the cholinergic vasodilator response due to sepsis. Taken together, these data demonstrated that statins reverse microvascular dysfunction and reduce neuroinflammation during sepsis, preventing the development of long-term cognitive decline.

Combined therapy with metformin and insulin attenuates systemic and hepatic alterations in a model of high-fat diet-/streptozotocin-induced diabetes.

In this study we have explored the pathogenesis of the hepatic alterations which occur in diabetes and the modulation of these complications by the combination of metformin adjunct treatment and insulin monotherapy. For this purpose, diabetic rats were treated with insulin (DM + Ins) or metformin plus insulin (DM + Met + Ins). Biochemical and cardiometabolic parameters were analysed by spectrophotometry. Intravital microscopy was used to study the hepatic microcirculation. In the liver tissue, real-time PCR was used to analyse oxidative stress enzymes, inflammatory markers and receptors for advanced glycation end products (AGE) (RAGE) gene expression. Lipid peroxidation was assessed by thiobarbituric acid reactive species (TBARs) analyses. AGE deposition and RAGE protein expression were studied by fluorescence spectrophotometry and Western blot respectively. Body weight, %HbA1c , urea, total proteins and oxidative stress parameters were found to be similarly improved by insulin or Met + Ins treatments. On the other hand, Met + Ins treatment showed a more pronounced effect on fasting blood glucose level than insulin monotherapy. Fructosamine, uric acid, creatinine, albumin and amylase levels and daily insulin dose requirements were found to be only improved by the combined Met + Ins treatment. Liver, renal and pancreatic dysfunction markers were found to be more positively affected by metformin adjunct therapy when compared to insulin treatment. Liver microcirculation damage was found to be completely protected by Met + Ins treatment, while insulin monotherapy showed no effect. Our results suggest that oxidative stress, microcirculatory damage and glycated proteins could be involved in the aetiology of liver disease due to diabetes. Additionally, metformin adjunct treatment improved systemic and liver injury in induced diabetes and showed a more pronounced effect than insulin monotherapy.

Statins decrease neuroinflammation and prevent cognitive impairment after cerebral malaria.

Cerebral malaria (CM) is the most severe manifestation of Plasmodium falciparum infection in children and non-immune adults. Previous work has documented a persistent cognitive impairment in children who survive an episode of CM that is mimicked in animal models of the disease. Potential therapeutic interventions for this complication have not been investigated, and are urgently needed. HMG-CoA reductase inhibitors (statins) are widely prescribed for cardiovascular diseases. In addition to their effects on the inhibition of cholesterol synthesis, statins have pleiotropic immunomodulatory activities. Here we tested if statins would prevent cognitive impairment in a murine model of cerebral malaria. Six days after infection with Plasmodium berghei ANKA (PbA) mice displayed clear signs of CM and were treated with chloroquine, or chloroquine and lovastatin. Intravital examination of pial vessels of infected animals demonstrated a decrease in functional capillary density and an increase in rolling and adhesion of leukocytes to inflamed endothelium that were reversed by treatment with lovastatin. In addition, oedema, ICAM-1, and CD11b mRNA levels were reduced in lovastatin-treated PbA-infected mice brains. Moreover, HMOX-1 mRNA levels are enhanced in lovastatin-treated healthy and infected brains. Oxidative stress and key inflammatory chemokines and cytokines were reduced to non-infected control levels in animals treated with lovastatin. Fifteen days post-infection cognitive dysfunction was detected by a battery of cognition tests in animals rescued from CM by chloroquine treatment. In contrast, it was absent in animals treated with lovastatin and chloroquine. The outcome was similar in experimental bacterial sepsis, suggesting that statins have neuroprotective effects in severe infectious syndromes in addition to CM. Statin treatment prevents neuroinflammation and blood brain barrier dysfunction in experimental CM and related conditions that are associated with cognitive sequelae, and may be a valuable adjuvant therapeutic agent for prevention of cognitive impairment in patients surviving an episode of CM.

Unveiling Lovastatin's Anti-Inflammatory Potential in Mouse's Brain during Acute Trypanosoma cruzi Infection.

Neurological commitment is a neglected manifestation of Chagas disease (CD). Meningoencephalitis mainly affects children and immunosuppressed patients, while stroke can occur with or without cardiac compromise. One of the possible causes of stroke development is microvascular commitment. Our group previously described that experimental Trypanossoma cruzi acute infection leads to cerebral microvasculopathy. This condition is characterized by decreased capillary density, increased leukocyte rolling and adhesion, and endothelial dysfunction. CD was discovered 114 years ago, and until today, only two drugs have been available for clinical treatment: benznidazole and nifurtimox. Both present a high cure rate for the acute phase (80%) and small cure rate for the chronic phase (20%). In addition, the high occurrence of side-effects, without proper medical follow-up, can result in treatment abandonment. Therefore, the search for new therapeutic schemes is necessary. Statins are drugs already used in the clinic that have several pleiotropic effects including endothelial function improvement, anti-inflammatory action, as well as trypanocidal effects, making them a potential alternative treatment for brain microvasculopathy in CD. Here, we investigate the effect of lovastatin (LOV) on brain microvasculopathy and inflammatory parameters. Swiss Webster mice were intraperitoneally inoculated with the Y strain of T. cruzi. Treatment with lovastatin (20 mg/kg/day) was initiated 24 h after the infection and continued for 14 consecutive days. We observed that LOV treatment did not affect parasitemia, brain microcirculation alterations, or the reduction in cerebral blood flow caused by T. cruzi infection. Also, LOV did not prevent the increased number of CD3+ cells and eNOS levels in the T. cruzi-infected brain. No alterations were observed on VCAM-1 and MCP-1 expressions, neither caused by infection nor LOV treatment. However, LOV prevented the increase in F4/80+ cells and ICAM-1 levels in the brain caused by acute infection with T. cruzi. These results suggest an anti-inflammatory activity of LOV, but more studies are needed to elucidate the role of LOV in CD acute infection.

Cardiac microvascular rarefaction in hyperthyroidism-induced left ventricle dysfunction.

OBJECTIVE: The pathophysiology underlying hyperthyroidism-induced left ventricle (LV) dysfunction and hypertrophy directly involves the heart and indirectly involves the neuroendocrine systems. The effects of hyperthyroidism on the microcirculation are still controversial in experimental models. We investigated the effects of hyperthyroidism on the cardiac function and microcirculation of an experimental rat model. METHODS: Male Wistar rats (170-250 g) were divided into two groups: the euthyroid group (n = 10), which was treated with 0.9% saline solution, and the hyperthyroid group (n = 10), which was treated with l-thyroxine (600 µg/kg/day, i.p.) during 14 days. An echocardiographic study was performed to evaluate the alterations in cardiac function, structure and geometry. The structural capillary density and the expression of angiotensin II AT1 receptor in the LV were analyzed using histochemistry and immunohistochemistry, respectively. RESULTS: Hyperthyroidism was found to induce profound cardiovascular alterations, such as systolic hypertension, tachycardia, LV dysfunction, cardiac hypertrophy, and myocardial fibrosis. This study demonstrates the existence of structural capillary rarefaction and the down-regulation of the cardiac angiotensin II AT1 receptor in the myocardium of hyperthyroid rats in comparison with euthyroid rats. CONCLUSIONS: Microvascular rarefaction may be involved in the pathophysiology of hyperthyroidism-induced cardiovascular alterations.

Antihypertensive treatment improves microvascular rarefaction and reactivity in low-risk hypertensive individuals.

OBJECTIVE: To test whether long-term antihypertensive treatment with metoprolol succinate (a ß1-adrenoceptor blocker) or olmesartan medoxomil (an angiotensin II AT1-receptor blocker) reverses microvascular dysfunction in hypertensive patients. METHODS: This study included 44 hypertensive outpatients and 20 age and sex-matched healthy controls. We used skin capillaroscopy to measure capillary density and recruitment at rest and during PORH. Endothelium-dependent vasodilation of skin microcirculation was evaluated with a LDPM system in combination with ACh iontophoresis, PORH, and LTH. RESULTS: Pretreatment capillary density in hypertensive patients was significantly reduced compared with controls (71.3 ± 1.5 vs. 80.6 ± 1.8 cap/mm²; p < 0.001), as was PORH (71.7 ± 1.5 vs. 79.5 ± 2.6 cap/mm²; p < 0.05). After treatment for six months, capillary density increased to 75.4 ± 1.1 cap/mm² (p < 0.01) at rest and 76.8 ± 1.1 cap/mm² during PORH. During LTH, CVC in perfusion units (PU)/mmHg was similar in patients (1.71 [1.31-2.12]) and controls (1.60 [1.12-1.91]) and increased significantly (1.82 [1.30-2.20]) after treatment. Maximal CVC during PORH was reduced in hypertensive patients (0.30 [0.22-0.39]) compared to controls (0.39 [0.31-0.49], p < 0.001) and increased (0.41 [0.29-0.51], p < 0.001) after treatment. CONCLUSIONS: Capillary rarefaction and microvascular endothelial dysfunction in hypertensive patients responded favorably to long-term pharmacological treatment.

Blockade of the renin-angiotensin system improves cerebral microcirculatory perfusion in diabetic hypertensive rats.

We examined the functional and structural microcirculatory alterations in the brain, skeletal muscle and myocardium of non-diabetic spontaneously hypertensive rats (SHR) and diabetic SHR (D-SHR), as well as the effects of long-term treatment with the angiotensin AT1-receptor antagonist olmesartan and the angiotensin-converting enzyme inhibitor enalapril. Diabetes was experimentally induced by a combination of a high-fat diet with a single low dose of streptozotocin (35 mg/kg, intraperitoneal injection). D-SHR were orally administered with olmesartan (5 mg/kg/day), enalapril (10 mg/kg/day) or vehicle for 28 days, and compared with vehicle-treated non-diabetic SHR or normotensive non-diabetic Wistar-Kyoto rats. The cerebral and skeletal muscle functional capillary density of pentobarbital-anesthetized rats was assessed using intravital fluorescence videomicroscopy. Chronic treatment with olmesartan or enalapril significantly lowered blood pressure and reversed brain functional capillary rarefaction. Brain oxidative stress was reduced to non-diabetic control levels in animals treated with olmesartan or enalapril. Histochemical analysis of the structural capillary density showed that both olmesartan and enalapril increased the capillary-to-fiber ratio in skeletal muscle and the capillary-to-fiber volume density in the left ventricle. Olmesartan and enalapril also prevented collagen deposition and the increase in cardiomyocyte diameter in the left ventricle. Our results suggest that the association between hypertension and diabetes results in microvascular alterations in the brain, skeletal muscle and myocardium that can be prevented by chronic blockade of the renin-angiotensin system.

Long-term assessment of systemic microcirculatory function and plasma cytokines after coronavirus disease 2019 (COVID-19).

Systemic microvascular dysfunction has been shown to be present in COVID-19, and serum cytokines are known to be involved in the regulation of vascular function. We sought to evaluate systemic microvascular endothelial function, with laser doppler perfusion monitoring (LDPM), and plasma levels of cytokines after acute COVID-19. Individuals admitted to a Cardiology hospital with acute COVID-19 and followed for 12-15 months after recovery underwent noninvasive evaluation of systemic endothelium-dependent microvascular reactivity by cutaneous LDPM with local thermal hyperemia (LTH). A multiplex biometric immunoassay panel was used to assess 48 serum cytokines and chemokines. Twenty patients and 14 control volunteers were enrolled. The areas under the curves of vasodilation induced by LTH were significantly increased after recovery (P=0.009) and were not different from values obtained in healthy volunteers (P = 0.85). The peak microvascular flow during LTH did also significantly increase (P = 0.02), and was not different form values obtained in healthy volunteers (P = 0.55). Several cytokines displayed significantly reduced serum concentrations after recovery from COVID-19. In conclusion, endothelium-dependent systemic microvascular reactivity improved after recovery from COVID-19 in patients with cardiovascular diseases, in parallel with a reduction in the levels of several serum cytokines and chemokines involved in the regulation of vascular function and inflammation.

Effect of benznidazole on cerebral microcirculation during acute Trypanosoma cruzi infection in mice.

Central nervous system alterations was described in Chagas disease in both human and experimental models, leading to meningoencephalitis, stroke and cognitive impairment. Recently, our group demonstrated that acute infection by Trypanossoma cruzi leads to cerebral microvasculophaty in mice with endothelial dysfunction, capillary rarefaction, increased rolling and leukocyte adhesion. Only benznidazole and nifurtimox are available for clinical treatment, they have an efficiency of 80% in the acute phase and less than 20% in chronic phase. However, the effect of these drugs on brain microcirculation has not yet been evaluated. We hypothesized that early treatment with benznidazole could protect brain microcirculation during acute experimental Chagas disease. Swiss Webster mice were inoculated with 104 trypomastigotes forms of T. cruzi, and after 24 h they were treated with 50 or 100 mg/kg/day of benznidazole for 14 consecutive days. In untreated infected mice, we observed cerebral microvascular rarefaction, increase in leukocyte rolling and adhesion, reduced cerebral blood flow, and increased CD3+ and F4-80+ cells in brain tissue. Early treatment with benznidazole at 100 mg/kg/day and 50 mg/kg/day prevented the occurrence of the alterations mentioned. Here, we show that BZ is able to protect the microcirculation and reduced brain inflammation in acute experimental Chagas disease.

Obesity-Related Inflammation and Endothelial Dysfunction in COVID-19: Impact on Disease Severity.

The coronavirus disease 2019 (COVID-19) pandemic has put into evidence another pandemic - obesity. Currently, several studies have documented the association between obesity and COVID-19 severity. The mechanisms underlying the increased risk of complications and mortality in obese patients with COVID-19 are of diverse nature. Inflammation plays a central role in obesity. Metabolic alterations seen in obese patients are related to an inflammatory response, and several studies report elevated levels of circulating inflammatory cytokines in obese patients. Also, deregulated expression of adipokines, such as leptin and resistin, increase the expression of vascular adhesion molecule 1 and intercellular adhesion molecule 1 that contribute to increased vascular leukocyte adhesiveness and additional oxidative stress. Additionally, it is now recognized that the chronic impairment of systemic vascular endothelial function in patients with cardiovascular and metabolic disorders, including obesity, when intensified by the detrimental effects of SARS-CoV-2 over the endothelium, may explain their worse outcomes in COVID-19. In fact, vascular endothelial dysfunction may contribute to a unfavorable response of the endothelium to the infection by SARS-CoV-2, whereas alterations in cardiac structure and function and the prothrombotic environment in obesity may also provide a link to the increased cardiovascular events in these patients.

Mesenchymal stromal cells protect against vascular damage and depression-like behavior in mice surviving cerebral malaria.

BACKGROUND: Malaria is one of the most critical global infectious diseases. Severe systemic inflammatory diseases, such as cerebral malaria, lead to the development of cognitive and behavioral alterations, such as learning disabilities and loss of memory capacity, as well as increased anxiety and depression. The consequences are profound and usually contribute to reduce the patient's quality of life. There are no therapies to treat the neurological sequelae of cerebral malaria. Mesenchymal stromal cells (MSCs) may be an alternative, since they have been used as therapy for neurodegenerative diseases and traumatic lesions of the central nervous system. So far, no study has investigated the effects of MSC therapy on the blood-brain barrier, leukocyte rolling and adherence in the brain, and depression like-behavior in experimental cerebral malaria. METHODS: Male C57BL/6 mice were infected with Plasmodium berghei ANKA (PbA, 1 × 106 PbA-parasitized red blood cells, intraperitoneally). At day 6, PbA-infected animals received chloroquine (25 mg/kg orally for seven consecutive days) as the antimalarial treatment and were then randomized to receive MSCs (1 × 105 cells in 0.05 ml of saline/mouse) or saline (0.05 ml) intravenously. Parasitemia, clinical score, and survival rate were analyzed throughout the experiments. Evans blue assay was performed at 6, 7, and 15 days post-infection (dpi). Behavioral tests were performed at 5 and 15 dpi. Intravital microscopy experiments and brain-derived neurotrophic factor (BDNF) protein expression analyses were performed at 7 dpi, whereas inflammatory mediators were measured at 15 dpi. In vitro, endothelial cells were used to evaluate the effects of conditioned media derived from MSCs (CMMSC) on cell viability by lactate dehydrogenase (LDH) release. RESULTS: PbA-infected mice presented increased parasitemia, adherent leukocytes, blood-brain barrier permeability, and reduced BDNF protein levels, as well as depression-like behavior. MSCs mitigated behavioral alterations, restored BDNF and transforming growth factor (TGF)-ß protein levels, and reduced blood-brain barrier dysfunction and leukocyte adhesion in the brain microvasculature. In a cultured endothelial cell line stimulated with heme, CMMSC reduced LDH release, suggesting a paracrine mechanism of action. CONCLUSION: A single dose of MSCs as adjuvant therapy protected against vascular damage and improved depression-like behavior in mice that survived experimental cerebral malaria.