Blockade of sympathetic ganglia improves vascular dysfunction in septic shock.

Sepsis/septic shock activates the sympathetic nervous system (SNS) to deal with the infection stress. However, an imbalanced or maladaptive response due to excessive or uncontrolled activation characterizes autonomic dysfunction. Our hypothesis was that reducing this excessive activation of the autonomic nervous system would impact positively in sepsis. Using ganglionic blockers as a pharmacological approach, the main aim of the present report was to assess the role of ganglionic transmission in the vascular dysfunction associated with sepsis.Sepsis was induced in rats by cecal ligation and puncture (CLP). One hour after CLP surgery, rats were treated subcutaneously with hexamethonium (15 mg/kg; ganglionic blocker), pentolinium (5 mg/kg; a blocker with a higher selectivity for sympathetic ganglia compared to hexamethonium), or vehicle (PBS). Basal blood pressure and the response to adrenergic agonists were evaluated at 6 and 24 h after CLP surgery. Reactivity to vasoconstrictors, nitric oxide (NO) synthase 2 (NOS-2) expression, IL-1 and TNF plasma levels, and density of α1 adrenergic receptors were evaluated in the aorta 24 h after CLP.Septic shock resulted in hypotension and hyporesponsiveness to norepinephrine and phenylephrine, increased plasma cytokine levels and NOS-2 expression in the aorta, and decreased α1 receptor density in the same vessel. Pentolinium but not hexamethonium recovered responsiveness and α1 adrenergic receptor density in the aorta. Both blockers normalized the in vivo response to vasoconstrictors, and reduced plasma IL-1 and NOx levels and NOS-2 expression in the aorta.Blockade of ganglionic sympathetic transmission reduced the vascular dysfunction in experimental sepsis. This beneficial effect seems to be, at least in part, due to the preservation of α1 adrenergic receptor density and to reduced NOS-2 expression and may lead to adjuvant ways to treat human sepsis.

Pharmacokinetics, biodistribution, and in vivo toxicity of 7-nitroindazole loaded in pegylated and non-pegylated nanoemulsions in rats.

Sepsis is a life-threatening condition caused by a dysregulated host response to infection. The development of sepsis is associated with excessive nitric oxide (NO) production, which plays an important role in controlling vascular homeostasis. 7-nitroindazole (7-NI) is a selective inhibitor of neuronal nitric oxide synthase (NOS-1) with potential application for treating NO imbalance conditions. However, 7-NI exhibits a low aqueous solubility and a short plasma half-life. To circumvent these biopharmaceutical limitations, pegylated (NEPEG7NI) and non-pegylated nanoemulsions (NENPEG7NI) containing 7-NI were developed. This study evaluates the pharmacokinetic profiles and toxicological properties of 7-NI loaded into the nanoemulsions. After a single intravenous administration of the free drug and the nanoemulsions at a dose of 10 mg.kg-1 in Wistar rats, 7-NI was widely distributed in the organs. The pharmacokinetic parameters of Cmax, t1/2, and AUC0-t were significantly increased after administration of the NEPEG7NI, compared to both free 7-NI and NENPEG7NI (p < 0.05). No observable adverse effects were observed after administering the free 7-NI, NEPEG7NI, or NENPEG7NI in the animals after a single dose of up to 3.0 mg.kg-1. The results indicated that 7-NI-loaded nanoemulsions are safe, constituting a promising approach to treating sepsis.

Cannabinoid CB2 receptor agonist reduces local and systemic inflammation associated with pneumonia-induced sepsis in mice.

Sepsis is a severe condition secondary to dysregulated host response to infection leading to tissue damage and organ dysfunction. Cannabinoid CB2 receptor has modulatory effects on the immune response. Therefore, this study investigated the effects of a cannabinoid CB2 receptor agonist on the local and systemic inflammatory process associated with pneumonia-induced sepsis. Pneumonia-induced sepsis was induced in mice by intratracheal inoculation of Klebsiella pneumoniae. Tissue and bronchoalveolar lavage (BAL) were collected 6, 24, or 48 h after surgery. Mice were treated with CB2 agonist (AM1241, 0.3 and 3 mg/kg, i.p.) and several parameters of inflammation were evaluated 24 h after sepsis induction. Polymorphonuclear cell migration to the infectious focus peaked 24 h after pneumonia-induced sepsis induction in male and female animals. Septic male mice presented a significant reduction of cannabinoid CB2 receptor density in the lung tissue after 24 h, which was not observed in females. CB2 expression in BAL macrophages was also reduced in septic animals. Treatment of septic mice with AM1241 reduced cell migration, local infection, myeloperoxidase activity, protein extravasation, and NOS-2 expression in the lungs. In addition, the treatment reduced plasma IL-1ß, increased IL-10 and reduced the severity and mortality of septic animals. These results suggest that AM1241 promotes an interesting balance in the inflammatory response, maintaining lung function and preventing organ injury. Therefore, cannabinoid CB2 receptors are potential targets to control the excessive inflammatory process that occurs in severe conditions, and agonists of these receptors can be considered promising adjuvants in pneumonia-induced sepsis treatment.

Relationship between peripheral ischemic microvascular reserve, persistent hyperlactatemia, and its temporal dynamics in sepsis: a post hoc study.

OBJECTIVE: To measure the prognostic value of peripheral ischemic microvascular reserve in the context of persistent sepsis-induced hyperlactatemia and measure its influence on the temporal dynamics of lactate and the strength of association between these variables. METHODS: This post hoc analysis of the peripheral perfusion index/postocclusive reactive hyperemia trial, an observational cohort study that enrolled patients with sepsis who persisted with lactate levels ≥ 2mmol/L after fluid resuscitation (with or without shock). Peripheral ischemic microvascular reserve was evaluated using the association of the peripheral perfusion index and postocclusive reactive hyperemia techniques. The cutoff point of ∆ peripheral perfusion index peak values (%) defined the groups with low (≤ 62%) and high peripheral ischemic microvascular reserve (> 62%). RESULTS: A total of 108 consecutive patients with persistent sepsis-induced hyperlactatemia were studied. The high peripheral ischemic microvascular reserve group showed higher 28-day mortality than the low peripheral ischemic microvascular reserve group (p < 0.01). The temporal dynamics of lactate within the first 48 hours showed a rapid decrease in lactate levels in the low peripheral ischemic microvascular reserve group (p < 0.01). However, this result was not reproduced in the linear mixed effects model. A weak correlation between peripheral ischemic microvascular reserve (%) and lactate level (mmol/L) was observed within the first 24 hours (r = 0.23; p < 0.05). CONCLUSION: The prognostic value of high peripheral ischemic microvascular reserve was confirmed in the context of persistent sepsis-induced hyperlactatemia. Although there was a weak positive correlation between peripheral ischemic microvascular reserve value and lactate level within the first 24 hours of sepsis diagnosis, the low peripheral ischemic microvascular reserve group appeared to have a faster decrease in lactate over the 48 hours of follow-up.

Peripheral ischemic reserve in sepsis and septic shock as a new bedside prognostic enrichment tool: A Brazilian cohort study.

Microvascular dysfunctions are associated with poor prognosis in sepsis. However, the potential role of clinical assessment of peripheral ischemic microvascular reserve (PIMR), a parameter that characterizes the variation of peripheral perfusion index (PPI) after brief ischemia of the upper arm, as a tool to detect sepsis-induced microvascular dysfunction and for prognostic enrichment has not been established. To address this gap, this study investigated the association of high PIMR with mortality over time in patients with sepsis and its subgroups (with and without shock) and peripheral perfusion (capillary-refill time). This observational cohort study enrolled consecutive septic patients in four Intensive-care units. After fluid resuscitation, PIMR was evaluated using the oximetry-derived PPI and post-occlusive reactive hyperemia for two consecutive days in septic patients. Two hundred and twenty-six patients were included-117 (52%) in the low PIMR group and 109 (48%) in the high PIMR group. The study revealed differences in mortality between groups on the first day, which was higher in the high PIMR group (RR 1.25; 95% CI 1.00-1.55; p = 0.04) and maintained its prognostic significance after multivariate adjustment. Subsequently, this analysis was made for sepsis subgroups and showed significant differences in mortality only for the septic-shock subgroup, with was higher in the high PIMR group (RR 2.14; 95% CI 1.49-3.08; p = 0.01). The temporal ΔPPI peak values (%) analyses did not demonstrate maintenance of the predictive value over the first 48 h in either group (p > 0.05). A moderate positive correlation (r = 0.41) between ΔPPI peak (%) and capillary-refill time (s) was found within the first 24 hours of diagnosis (p < 0.001). In conclusion, detecting a high PIMR within 24 h appears to be a prognostic marker for mortality in sepsis. Furthermore, its potential as a prognostic enrichment tool seems to occur mainly in septic shock.

Role of nitric oxide on defensive behavior and long-term aversive learning induced by chemical stimulation of the dorsolateral periaqueductal gray matter.

The midbrain periaqueductal gray matter, especially the dorsolateral portion (dlPAG), coordinates immediate defensive responses (DR) to threats, but also ascends forebrain information for aversive learning. The synaptic dynamics in the dlPAG regulate the intensity and type of behavioral expression, as well as long-term processes such as memory acquisition, consolidation, and retrieval. Among several neurotransmitters and neural modulators, nitric oxide seems to play an important regulatory role in the immediate expression of DR, but it remains unclear if this gaseous on-demand neuromodulator contributes to aversive learning. Therefore, the role of nitric oxide in the dlPAG was investigated, during conditioning in an olfactory aversive task. The behavioral analysis consisted of freezing and crouch-sniffing in the conditioning day after glutamatergic NMDA agonist injection into the dlPAG. Two days later, rats were re-exposed to the odor cue and avoidance was measured. 7NI, a selective neuronal nitric oxide synthase inhibitor (40 and 100 nmol), injected before NMDA (50 pmol) impaired immediate DR and consequent aversive learning. The scavenging of extrasynaptic nitric oxide by C-PTIO (1 and 2 nmol) induced similar results. Moreover, spermine NONOate, a nitric oxide donor (5, 10, 20, 40, and 80 nmol), produced DR by itself, but only the low dose also promoted learning. The following experiments utilized a fluorescent probe, DAF-FM diacetate (5 µM), directly into the dlPAG, to quantify nitric oxide in the three previous experimental situations. Nitric oxide levels were increased after NMDA stimulation, decreased after 7NI, and increased after spermine NONOate, in line with alterations in defensive expression. Altogether, the results indicate that nitric oxide plays a modulatory and decisive role in the dlPAG regarding immediate DR and aversive learning.

Relationship between peripheral ischemic microvascular reserve, persistent hyperlactatemia, and its temporal dynamics in sepsis: a post hoc study / Relação entre reserva microvascular isquêmica periférica, hiperlactatemia persistente e sua dinâmica temporal na sepse: um estudo post hoc

ABSTRACT Objective: To measure the prognostic value of peripheral ischemic microvascular reserve in the context of persistent sepsis-induced hyperlactatemia and measure its influence on the temporal dynamics of lactate and the strength of association between these variables. Methods: This post hoc analysis of the peripheral perfusion index/postocclusive reactive hyperemia trial, an observational cohort study that enrolled patients with sepsis who persisted with lactate levels ≥ 2mmol/L after fluid resuscitation (with or without shock). Peripheral ischemic microvascular reserve was evaluated using the association of the peripheral perfusion index and postocclusive reactive hyperemia techniques. The cutoff point of ∆ peripheral perfusion index peak values (%) defined the groups with low (≤ 62%) and high peripheral ischemic microvascular reserve (> 62%). Results: A total of 108 consecutive patients with persistent sepsis-induced hyperlactatemia were studied. The high peripheral ischemic microvascular reserve group showed higher 28-day mortality than the low peripheral ischemic microvascular reserve group (p < 0.01). The temporal dynamics of lactate within the first 48 hours showed a rapid decrease in lactate levels in the low peripheral ischemic microvascular reserve group (p < 0.01). However, this result was not reproduced in the linear mixed effects model. A weak correlation between peripheral ischemic microvascular reserve (%) and lactate level (mmol/L) was observed within the first 24 hours (r = 0.23; p < 0.05). Conclusion: The prognostic value of high peripheral ischemic microvascular reserve was confirmed in the context of persistent sepsis-induced hyperlactatemia. Although there was a weak positive correlation between peripheral ischemic microvascular reserve value and lactate level within the first 24 hours of sepsis diagnosis, the low peripheral ischemic microvascular reserve group appeared to have a faster decrease in lactate over the 48 hours of follow-up.

RESUMO Objetivo: Avaliar o valor prognóstico da reserva microvascular isquêmica periférica no contexto da hiperlactatemia persistente induzida pela sepse, determinar sua influência na dinâmica temporal de lactato e analisar a força da associação entre essas variáveis. Métodos: Análise post hoc do estudo de índice de perfusão periférica/hiperemia reativa pós-oclusiva caracterizada por uma coorte observacional que incluiu pacientes com sepse que persistiram com níveis de lactato ≥ 2mmol/L após a ressuscitação volêmica (com ou sem choque). A reserva microvascular isquêmica periférica foi mensurada utilizando-se a associação dos métodos do índice de perfusão periférica e hiperemia reativa pós-oclusiva. O ponto de corte dos valores da ∆ índice de perfusão periférica de pico (%) definiu os grupos com baixa (≤ 62%) e alta (> 62%) reserva microvascular isquêmica periférica. Resultados: Estudaram-se 108 pacientes consecutivos com hiperlactatemia persistente induzida pela sepse. O grupo com alta reserva microvascular isquêmica periférica apresentou maior mortalidade aos 28 dias em relação ao grupo com baixa reserva microvascular isquêmica periférica (p < 0,01). A dinâmica temporal de lactato nas primeiras 48 horas mostrou redução rápida dos níveis de lactato no grupo com baixa reserva microvascular isquêmica periférica (p < 0,01). No entanto, esse resultado não foi reproduzido no modelo de efeitos mistos lineares. Observou-se fraca correlação (%) entre os valores da reserva microvascular isquêmica periférica e níveis de lactato (mmol/L) nas primeiras 24 horas (r = 0,23; p < 0,05). Conclusão: O valor prognóstico da alta reserva microvascular isquêmica periférica foi confirmado no contexto da hiperlactatemia persistente induzida por sepse. Embora tenha sido observada uma baixa correlação positiva entre os valores da reserva microvascular isquêmica periférica e os níveis de lactato nas primeiras 24 horas do diagnóstico de sepse, o grupo com baixa reserva microvascular isquêmica periférica pareceu apresentar redução mais rápida do lactato nas 48 horas de seguimento.

The role of nitric oxide in sepsis-associated kidney injury.

Sepsis is one of the leading causes of acute kidney injury (AKI), and several mechanisms including microcirculatory alterations, oxidative stress, and endothelial cell dysfunction are involved. Nitric oxide (NO) is one of the common elements to all these mechanisms. Although all three nitric oxide synthase (NOS) isoforms are constitutively expressed within the kidneys, they contribute in different ways to nitrergic signaling. While the endothelial (eNOS) and neuronal (nNOS) isoforms are likely to be the main sources of NO under basal conditions and participate in the regulation of renal hemodynamics, the inducible isoform (iNOS) is dramatically increased in conditions such as sepsis. The overexpression of iNOS in the renal cortex causes a shunting of blood to this region, with consequent medullary ischemia in sepsis. Differences in the vascular reactivity among different vascular beds may also help to explain renal failure in this condition. While most of the vessels present vasoplegia and do not respond to vasoconstrictors, renal microcirculation behaves differently from nonrenal vascular beds, displaying similar constrictor responses in control and septic conditions. The selective inhibition of iNOS, without affecting other isoforms, has been described as the ideal scenario. However, iNOS is also constitutively expressed in the kidneys and the NO produced by this isoform is important for immune defense. In this sense, instead of a direct iNOS inhibition, targeting the NO effectors such as guanylate cyclase, potassium channels, peroxynitrite, and S-nitrosothiols, may be a more interesting approach in sepsis-AKI and further investigation is warranted.

Dynamics of GRK2 in the kidney: a putative mechanism for sepsis-associated kidney injury.

Renal vascular reactivity to vasoconstrictors is preserved in sepsis in opposition to what happens in the systemic circulation. We studied whether this distinct behavior was related to α1 adrenergic receptor density, G protein-coupled receptor kinase 2 (GRK2) and the putative role of nitric oxide (NO). Sepsis was induced in female mice by cecal ligation and puncture (CLP). Wildtype mice were treated with prazosin 12 h after CLP or nitric oxide synthase 2 (NOS-2) inhibitor, 30 min before and 6 and 12 h after CLP. In vivo experiments and biochemistry assays were performed 24 h after CLP. Sepsis decreased the systemic mean arterial pressure (MAP) and the vascular reactivity to phenylephrine. Sepsis also reduced basal renal blood flow which was normalized by treatment with prazosin. Sepsis led to a substantial decrease in GRK2 level associated with an increase in α1 adrenergic receptor density in the kidney. The disappearance of renal GRK2 was prevented in NOS-2-KO mice or mice treated with 1400 W. Treatment of non-septic mice with an NO donor reduced GRK2 content in the kidney. Therefore, our results show that an NO-dependent reduction in GRK2 level in the kidney leads to the maintenance of a normal α1 adrenergic receptor density. The preservation of the density and/or functionality of this receptor in the kidney together with a higher vasoconstrictor tonus in sepsis lead to vasoconstriction. Thus, the increased concentration of vasoconstrictor mediators together with the preservation (and even increase) of the response to them may help to explain sepsis-induced acute kidney injury.

Response surface methodology (RSM) to evaluate both the extraction of triterpenes and sterols from jackfruit seed with supercritical CO2 and the biological activity of the extracts.

Jackfruit seeds are an underestimate residue having important biological activity such as anti-inflammatory, cytotoxicity and antimicrobial effects. However few researches have been done for this material using alternative extraction technologies, so this study aimed to evaluate the extraction of triterpenes and sterols from jackfruit seed by applying high- and low-pressure techniques. Response surface methodology (RSM) was used to determine the best conditions of pressure, temperature and CO2 flow rate for extraction with supercritical CO2. The yield and profile of these compounds were compared with the low pressure technique, which was considered as a reference. In vitro biological tests of anti-inflammatory activity and cytotoxicity in L929 and RAW 264.7 cells were also performed. The best extraction conditions in SFE for sterols were 40 °C/20 MPa/4 mL min-1 (0.832 ± 0.007 mgSR g-1 sample) and 40 °C/20 MPa/3 mL min-1 (0.800 ± 0.009 mgSR g-1 sample), for triterpenes were 50 °C/12 MPa/4 mL min-1 (1.501 ± 0.004 mgTT g-1 sample) and 45 °C/9.3 MPa/3.5 mL min-1 (1.485 ± 0.004 mgTT g-1 sample). No cytotoxic activity was detected in L929 cells in the extracts obtained from ethanol up to concentration of 100 µg mL-1 of extract. The Pearson's coefficient indicated that the reduction in cell viability was related to the concentration of triterpenes. Anti-inflammatory assays showed that some extracts could inhibit the inflammatory action induced in RAW 264.7 cells at concentration of 30 µg mL-1 of extract. Our results justify the further exploration of these characteristics to obtain natural products for the pharmaceutical and food industries.

NO, ROS, RAS, and PVAT: More Than a Soup of Letters.

Perivascular adipose tissue (PVAT) has recently entered in the realm of cardiovascular diseases as a putative target for intervention. Notwithstanding its relevance, there is still a long way before the role of PVAT in physiology and pathology is fully understood. The general idea that PVAT anti-contractile effect is beneficial and its pro-contractile effect is harmful is being questioned by several reports. The role of some PVAT important products or systems such as nitric oxide (NO), reactive oxygen species (ROS), and RAS may vary depending on the context, disease, place of production, etc., which adds doubts on how mediators of PVAT anti- and pro-contractile effects are called to action and their final result. This short review will address some points regarding NO, ROS, and RAS in the beneficial and harmful roles of PVAT.

Angiotensin II receptor type 1 blockade improves hyporesponsiveness to vasopressors in septic shock.

Sepsis activates the renin-angiotensin system and the production of angiotensin II, which has a key role in the regulation of blood pressure through AT1 receptors. However, excessive activation of AT1 receptor is associated with deleterious effects. We investigated the consequences of a differential blockade of AT1 receptor caused by two doses of losartan (0.25 mg/kg or 15 mg/kg, s.c), a selective AT1 receptor antagonist on sepsis outcome. These doses reduced the effect of angiotensin II in normal rats by 30% and >90% 8 h after administration, respectively, but only the higher dose maintained its inhibitory effect (~70%) 24 h after injection. Sepsis was induced by cecal ligation and puncture (CLP). Losartan was injected 2 h after CLP and parameters were evaluated 6 and 24 h after CLP. Septic rats developed hypotension and hyporesponsiveness to vasoconstrictors, an intense inflammatory process and increase in plasma markers of organ dysfunction. The lower dose of losartan improved the vasoconstrictive response to phenylephrine and angiotensin II, reduced lung myeloperoxidase and prevented leukopenia 24 h after CLP, but it did not reduce NOS-2 expression, plasma IL-6 levels or organ injury parameters of septic rats. On the other hand, the higher dose of losartan worsened the response to vasoconstrictors, potentiated the hypotension and increased further levels of creatine, urea and lactate in septic rats. Therefore, an early and partial blockade of AT1 receptor with a low dose of losartan may counteract sepsis-induced refractoriness to vasoconstrictors thus providing an opportunity to improve the outcome of this condition.

The Therapeutic Value of Hydralazine in Reducing Inflammatory Response, Oxidative Stress, and Mortality in Animal Sepsis: Involvement of the PI3K/AKT Pathway.

ABSTRACT: Sepsis is an amplified systemic immune-inflammatory response produced by a microorganism, which involves activation of inflammatory cytokine signaling pathways and oxidative stress. A variety of studies have shown that hydralazine (HDZ) has potent antioxidant and anti-inflammatory proprieties. Therefore, we hypothesize that HDZ can improve the clinical outcome of sepsis. Thus, this work aimed to evaluate therapeutic value of HDZ in reducing inflammatory response, oxidative stress, and mortality in animal sepsis, and to investigate its possible mechanism of action. Sepsis was induced by the cecal ligation and puncture (CLP) method in Wistar rats. After surgery, the animals were randomly divided into three groups: sham, sepsis, and sepsis + HDZ (1 mg/kg, s.c.). All groups were monitored for 48 h to assess survival rate, and clinical, hemodynamic, biochemical, and cellular parameters. After euthanasia, blood, spleen, liver, and kidneys were collected for analysis. Blood serum cytokines, tissue myeloperoxidase (MPO) activity, and oxidative stress parameters were assessed. Involvement of the PI3K/Akt pathway was also investigated. Sepsis was successfully induced by the CLP technique. HDZ treatment increased the survival rate (from 50% to 90%), improved glycemia control, reduced the clinical severity sepsis and mean arterial pressure; and prevented increased MPO activity, TNF-α, IL-1ß, IL-10 levels, and oxidative damage markers. Additionally, HDZ significantly prevented the increase of Akt activation in the liver and kidney. HDZ largely mitigated the effects of sepsis by suppressing inflammatory and antioxidant responses via the PI3K/Akt pathway. These findings provide evidence that HDZ can be a new therapeutic alternative for treating sepsis.

Aerobic exercise ameliorates survival, clinical score, lung inflammation, DNA and protein damage in septic mice.

BACKGROUND AND OBJECTIVE: Sepsis is a potentially deadly organic dysfunction, and one of the main causes of mortality in intensive care units (ICU). Aerobic exercise (AE) is a preventive intervention in the establishment of inflammatory conditions, such as chronic lung diseases, but its effects on sepsis remain unclear. Therefore, this study aimed to evaluate the effects of AE on health condition, mortality, inflammation, and oxidative damage in an experimental model of pneumosepsis induced by Klebsiella pneumoniae (K.p). METHODS: Animals were randomly allocated to Control; Exercise (EXE); Pneumosepsis (PS) or Exercise + Pneumosepsis (EPS) groups. Exercised animals were submitted to treadmill exercise for 2 weeks, 30 min/day, prior to pneumosepsis induced by K.p tracheal instillation. RESULTS: PS produced a striking decrease in the health condition leading to massive death (85%). AE protected mice, as evidenced by better clinical scores and increased survival (70%). AE alleviated sickness behavior in EPS mice as evaluated in the open field test, and inflammation (nitrite + nitrate, TNF-α and IL-1ß levels) in broncoalveolar fluid. Catalase activity, oxidative damage to proteins and DNA was increased by sepsis and prevented by exercise. CONCLUSION: Overall, the beneficial effects of exercise in septic animals encompassed a markedly improved clinical score and decreased mortality, along with lower inflammation markers, less DNA and protein damage, as well as preserved antioxidant enzyme activity. Neural network risk analysis revealed exercise had a considerable effect on the overall health condition of septic mice.

Perivascular adipose tissue phenotype and sepsis vascular dysfunction: Differential contribution of NO, ROS and beta 3-adrenergic receptor.

AIMS: Vascular dysfunction plays a key role in sepsis but the role of perivascular adipose tissue (PVAT) in this condition is relatively unknown. MAIN METHODS: Sepsis was induced by cecal ligation and puncture (CLP). The responses of the aorta and superior mesenteric artery to norepinephrine in the presence or absence of PVAT were evaluated. Fluorescent probes measured the production of nitric oxide (NO) and reactive oxygen species (ROS). NO synthases (NOS) and ß3-adrenoceptor expression were detected by immunofluorescence and S-nitrosylation by the biotin switch assay. KEY FINDINGS: Aorta and superior mesenteric arteries from septic animals with intact PVAT showed a worsened response to the vasoconstrictor compared to vessels without PVAT. PVAT from the aorta (APVAT) produced NO and ROS whereas PVAT from the superior mesenteric artery (MPVAT) produced only ROS. Septic APVAT exhibited a higher density of NOS-1 and NOS-3. S-nitrosylation was found in APVAT. Donor (PVAT obtained from normal or septic rats):Host (normal vessel without PVAT) experiments showed that L-NAME, ODQ and ß3-adrenergic receptor antagonist blocked the septic APVAT anti-contractile effect. None of these compounds affected MPVAT; tempol, but not apocynin, blocked its anti-contractile effect. SIGNIFICANCE: PVAT contributes to the anti-contractile effect in the aorta and mesenteric artery of septic rats through different pathways. ß3-Adrenergic receptor and NO appear to be key mediators of this effect in APVAT, but not in MPVAT where ROS seem to be a relevant mediator. Therefore, PVAT is a relevant player of sepsis vascular dysfunction.

The apoptosis clearance signal phosphatidylserine inhibits leukocyte migration and promotes inflammation resolution in vivo.

Previous studies have shown that phagocytosis of apoptotic cells can tune the macrophage phenotype and trigger the resolution of inflammation. This mechanism is largely dependent on the recognition of phosphatidylserine (PS) residues on the outer membrane of dying cells. Therefore, we sought to assess the effects of PS-containing liposomes (mimics of apoptotic cells) on the leukocyte migration profile during the inflammatory process in vivo. Inflammation was induced by carrageenan injection into air pouches created on the dorsal region of mice, as this model enables convenient access to the exudates for further investigation. Mice were treated with PBS, PS-containing or phosphatidylcholine (PC)-containing liposomes (10, 30 or 100 mg/kg intraperitoneally [i.p.]). Starting 8 h after carrageenan injection, the level of leukocyte infiltration was monitored over three days. The PS-containing, but not PC-containing, liposomes reduced the polymorphonuclear (PMN) and mononuclear (MN) leukocyte influx into the inflamed pouches in a dose-dependent fashion. Most notably, these effects could also be adoptively transferred; that is, they were also found in mice injected with a liposome-free peritoneal lavage obtained from the mice that had received the intraperitoneal PS-liposome treatment. The effect of treatment with the PS-induced soluble mediators (PS-ISMs) was found to be dependent on the presence of peritoneal macrophages and was susceptible to heat, trypsin degradation, and cycloheximide treatment. The PS-containing liposomes promoted the reduction of PMN leukocyte influx by triggering the release of anti-inflammatory autacoids with a proteinaceous nature that were produced de novo after PS exposure.

Corticoid-like anti-inflammatory effect of Vochysia bifalcata Warm.: Preclinical evidence of efficacy and safety.

ETHNOPHARMACOLOGICAL RELEVANCE: Vochysia bifalcata is a Brazilian native tree commonly used for economic purpose in the reforestation and in the manufacture of products. However, the potential usage of other parts of the plant is usually wasted. Besides, other species of Vochysia are well known for its anti-inflammatory action. AIM OF THE STUDY: In this study we evaluate the possible anti-inflammatory activity of the hydroethanolic extract from the leaves of V. bifalcata in models of mice skin inflammation. MATERIALS AND METHODS: Effects of V. bifalcata were evaluated in croton oil-induced acute and chronic skin inflammation. The role of glucocorticoid receptors in the extract effect was assessed by using a glucocorticoid receptor antagonist and by a specific binding assay. Possible adverse effects were evaluated after multiple treatments with the extract in a skin atrophy model. RESULTS: Topical application of V. bifalcata reduced ear edema formation, cell infiltration and interleukin (IL)-6 and tumor necrosis factor (TNF)-α levels. In the chronic model, besides edema formation and cell infiltration, the extract inhibited epidermal hyperproliferation and Proliferating Cell Nuclear Antigen expression. V. bifalcata seems to act by biding to corticoid receptors, however it did not induce corticoid related undesirable effects. CONCLUSION: Hydroethanolic extract from leaves of V. bifalcata could be an interesting tool in the search for new anti-inflammatory and antiproliferative agents for the treatment of skin disorders.

Bradykinin increases BP in endotoxemic rat: functional and biochemical evidence of angiotensin II AT1 /bradykinin B2 receptor heterodimerization.

BACKGROUND AND PURPOSE: Bradykinin may induce vasoconstriction in selected vessels or under specific experimental conditions. We hypothesized that inflammatory stimuli, such as endotoxin challenge, may induce the dimerization of AT1 /B2 receptors, altering the vascular effects of bradykinin. EXPERIMENTAL APPROACH: Wistar rats received LPS (1 mg·kg-1 , i.p.) and were anaesthetized for assessment of BP. Mesenteric resistance arteries were used in organ baths and subjected to co-immunoprecipitation and Western blot analyses. KEY RESULTS: At 24 and 48 hr after LPS, bradykinin-induced hypotension was followed by a sustained increase in BP, which was not found in non-endotoxemic animals. The B2 receptor antagonist Hoe-140 fully blocked the responses to bradykinin. The pressor effect of bradykinin was not prevented by prazosin, an α1 -adrenoceptor antagonist, but it was inhibited by the AT1 receptor antagonist losartan or the Rho-kinase inhibitor Y-27632. Endotoxemic rats also displayed enhanced pressor responses to angiotensin II, which were blocked by Hoe-140. Co-immunoprecipitation isolated using anti-B2 or anti-AT1 receptor antibodies showed that resistance arteries presented augmented levels of the AT1 /B2 receptor complexes at 24 hr after LPS injection. The presence of AT1 /B2 receptor heterodimers did correlate with the development of losartan-sensitive contractile responses to bradykinin and potentiation of angiotensin II-induced contraction, which was prevented by Hoe-140. CONCLUSIONS AND IMPLICATIONS: Endotoxin challenge is a stimulus for AT1 /B2 receptor heterodimerization in native vessels and shifts the B2 receptor-dependent vascular effect of bradykinin to a more complex pathway, which also depends on AT1 receptors and their intracellular signalling pathways.

Cyclodextrin based nanosponge of norfloxacin: Intestinal permeation enhancement and improved antibacterial activity.

Nanosponges are a novel class of hyperbranched cyclodextrin-based nanostructures that exhibits remarkable potential as a drug host system for the improvement in biopharmaceutical properties. This work aims the development of cyclodextrin-based nanosponge of norfloxacin to improve its physicochemical characteristics. ß-cyclodextrin was used as base and diphenyl carbonate as crosslinker agent at different proportions to produce nanosponges that were evaluated by in vitro and in vivo techniques. The proportion cyclodextrin:crosslinker 1:2 M/M was chosen due to its higher encapsulation efficiency (80%), revealing an average diameter size of 40 nm with zeta potential of -19 mV. Norfloxacin-loaded nanosponges exhibited higher passage of norfloxacin in comparison to norfloxacin drug alone by Ussing chamber method. The nanosponge formulation also revealed a mucoadhesive property that could increase norfloxacin absorption thus improving its antibiotic activity in an in vivo sepsis model. Therefore, nanosponges may be suitable carrier of norfloxacin to maximize and facilitate oral absorption.

7-nitroindazol-loaded nanoemulsions: Preparation, characterization and its improved inhibitory effect on nitric oxide synthase-1.

Nitric oxide (NO) participates in several physiological processes such as maintenance of blood pressure, host defense, neurotransmission, inhibition of platelet aggregation and learning and memory. NO is also involved in several diseases or dysfunctions in the cardiovascular, central nervous and musculoskeletal systems. NO also has been shown to be a major player in sepsis. NOS-1-derived NO has been shown to be a relevant species in physiology but also is an important element in pathology. There exist some NOS-1 inhibitors and among of them, 7-nitroindazole has been used for its in vivo selectivity. However, 7-NI has a very short half-life (∼2 h) and a poor water solubility. In this study, we describe the preparation and characterization of 7-NI-loaded nanoemulsions (NE7-NI). The chemical stability of 7-NI was greatly increased and the drug release rate could be controlled after nanoemulsification. NE7-NI reduced NO production in a long-lasting manner in vascular smooth muscle cells and skeletal muscle, without cytotoxicity. Our results evidenced that nanoemulsification approach increases the effective action time of 7-NI, rendering a suitable dosage form, which may be an interesting tool to study the role of NOS-1 in physiology and disease.