Diminazene aceturate attenuates systemic inflammation via microbiota gut-5-HT brain-spleen sympathetic axis in male mice.

The sympathetic arm of the inflammatory reflex is the efferent pathway through which the central nervous system (CNS) can control peripheral immune responses. Diminazene aceturate (DIZE) is an antiparasitic drug that has been reported to exert protective effects on various experimental models of inflammation. However, the pathways by which DIZE promotes a protective immunomodulatory effects still need to be well established, and no studies demonstrate the capacity of DIZE to modulate a neural reflex to control inflammation. C57BL/6 male mice received intraperitoneal administration of DIZE (2 mg/Kg) followed by lipopolysaccharide (LPS, 5 mg/Kg, i.p.). Endotoxemic animals showed hyperresponsiveness to inflammatory signals, while those treated with DIZE promoted the activation of the inflammatory reflex to attenuate the inflammatory response during endotoxemia. The unilateral cervical vagotomy did not affect the anti-inflammatory effect of DIZE in the spleen and serum. At the same time, splenic denervation attenuated tumor necrosis factor (TNF) synthesis in the spleen and serum. Using broad-spectrum antibiotics for two weeks showed that LPS modulated the microbiota to induce a pro-inflammatory profile in the intestine and reduced the serum concentration of tryptophan and serotonin (5-HT), while DIZE restored serum tryptophan and increased the hypothalamic 5-HT levels. Furthermore, the treatment with 4-Chloro-DL-phenylalanine (pcpa, an inhibitor of 5-HT synthesis) abolished the anti-inflammatory effects of the DIZE in the spleen. Our results indicate that DIZE promotes microbiota modulation to increase central 5-HT levels and activates the efferent sympathetic arm of the inflammatory reflex to control splenic TNF production in endotoxemic mice.

In vitro and in vivo antitrypanosomal activity of the fresh leaves of Ranunculus Multifidus Forsk and its major compound anemonin against Trypanosoma congolense field isolate.

BACKGROUND: Animal trypanosomiasis is a major livestock problem due to its socioeconomic impacts in tropical countries. Currently used trypanocides are toxic, expensive, and the parasites have developed resistance to the existing drugs, which calls for an urgent need of new effective and safe chemotherapeutic agents from alternative sources such as medicinal plants. In Ethiopian traditional medicine fresh leaves of Ranunculus multifidus Forsk, are used for the treatment of animal trypanosomiasis. The present study aimed to evaluate the antitrypanosomal activity of the fresh leaves of R. multifidus and its major compound anemonin against Trypanosoma congolense field isolate. METHODS: Fresh leaves of R. multifidus were extracted by maceration with 80% methanol and hydro-distillation to obtain the corresponding extracts. Anemonin was isolated from the hydro-distilled extract by preparative TLC. For the in vitro assay, 0.1, 0.4, 2 and 4 mg/ml of the test substances were incubated with parasites and cessation or drop in motility of the parasites was monitored for a total duration of 1 h. In the in vivo assay, the test substances were administered intraperitoneally daily for 7 days to mice infected with Trypanosoma congolense. Diminazene aceturate and 1% dimethylsulfoxide (DMSO) were used as positive and negative controls, respectively. RESULTS: Both extracts showed antitrypanosomal activity although the hydro-distilled extract demonstrated superior activity compared to the hydroalcoholic extract. At a concentration of 4 mg/ml, the hydro-distilled extract drastically reduced motility of trypanosomes within 20 min. Similarly, anemonin at the same concentration completely immobilized trypanosomes within 5 min of incubation, while diminazene aceturate (28.00 mg/kg/day) immobilized the parasites within 10 min. In the in vivo antitrypanosomal assay, anemonin eliminates parasites at all the tested doses (8.75, 17.00 and 35.00 mg/kg/day) and prevented relapse, while in diminazene aceturate-treated mice the parasites reappeared on days 12 to 14. CONCLUSIONS: The current study demonstrated that the fresh leaves of R. multifidus possess genuine antitrypanosomal activity supporting the use of the plant for the treatment of animal trypanosomiasis in traditional medicine. Furthermore, anemonin appears to be responsible for the activity suggesting its potential as a scaffold for the development of safe and cost effective antitrypanosomal agent.

Cardioprotective effects of losartan and diminazene against the ischemia/reperfusion injury in hyperthyroidism rats.

Hyperthyroidism is a condition where the thyroid gland produces high levels of thyroid hormone. Heart diseases are one of the main complications of hyperthyroidism. Several studies have shown that losartan (LOS) and diminazene aceturate (DIZE) possess cardioprotection effects against cardiac hypertrophy, ischemic heart disease, and heart failure. The research aimed to investigate the cardioprotection of LOS, DIZE, and their combination in the case of levothyroxine (LT4)-induced cardiomyopathy in rats. Hyperthyroidism was induced by LT4 in drinking water (12 mg/L) for 28 days. LOS (10 mg/kg, orally) and/or DIZE (15 mg/kg, subcutaneously) were administrated in rats with hyperthyroidism for 28 days. Decreased serum creatine kinase myoglobin and lactate dehydrogenase levels and cardiac hypertrophy by DIZE and combination therapy in hyperthyroidism rats have been reported. Cardiac hemodynamic findings showed that DIZE and its combination with LOS decreased the LT4-mediated left ventricular developed pressure (LVDP), rate pressure product (RPP), and RPP recovery percentage. Elevated cardiac oxidative stress and inflammation were confirmed by decreasing cardiac superoxide dismutase (SOD) activity and increasing the total oxidative stress and tumor necrosis factor-alpha (TNF-α) levels. SOD activity and TNF-α level were reversed by LOS and DIZE administration, respectively. Generally, DIZE and combination therapy with LOS improved cardiac dysfunction caused by hyperthyroidism in rats, whereas LOS alone has not been able to effectively respond to this dysfunction.

Diminazene resistance in Trypanosoma congolense is not caused by reduced transport capacity but associated with reduced mitochondrial membrane potential.

Trypanosoma congolense is a principal agent causing livestock trypanosomiasis in Africa, costing developing economies billions of dollars and undermining food security. Only the diamidine diminazene and the phenanthridine isometamidium are regularly used, and resistance is widespread but poorly understood. We induced stable diminazene resistance in T. congolense strain IL3000 in vitro. There was no cross-resistance with the phenanthridine drugs, melaminophenyl arsenicals, oxaborole trypanocides, or with diamidine trypanocides, except the close analogs DB829 and DB75. Fluorescence microscopy showed that accumulation of DB75 was inhibited by folate. Uptake of [3 H]-diminazene was slow with low affinity and partly but reciprocally inhibited by folate and by competing diamidines. Expression of T. congolense folate transporters in diminazene-resistant Trypanosoma brucei brucei significantly sensitized the cells to diminazene and DB829, but not to oxaborole AN7973. However, [3 H]-diminazene transport studies, whole-genome sequencing, and RNA-seq found no major changes in diminazene uptake, folate transporter sequence, or expression. Instead, all resistant clones displayed a moderate reduction in the mitochondrial membrane potential Ψm. We conclude that diminazene uptake in T. congolense proceed via multiple low affinity mechanisms including folate transporters; while resistance is associated with a reduction in Ψm it is unclear whether this is the primary cause of the resistance.

Inhibition of endoplasmic reticulum stress combined with activation of angiotensin-converting enzyme 2: novel approach for the prevention of endothelial dysfunction in type 1 diabetic rats.

Persistent hyperglycemia in type 1 diabetes triggers numerous signaling pathways, which may prove deleterious to the endothelium. As hyperglycemia damages the endothelial layer via multiple signaling pathways, including enhanced oxidative stress, downregulation of angiotensin-converting enzyme 2 signaling, and exacerbation of endoplasmic reticulum (ER) stress, it becomes difficult to prevent injury using monotherapy. Thus, the present study was conceived to evaluate the combined effect of ER stress inhibition along with angiotensin-converting enzyme 2 activation, two major contributors to hyperglycemia-induced endothelial dysfunction, in preventing endothelial dysfunction associated with type 1 diabetes. Streptozotocin-induced diabetic animals were treated with either diminazene aceturate (5 mg·kg-1 per day, p.o.) or tauroursodeoxycholic acid, sodium salt (200 mg·kg-1 per day i.p.), or both for 4 weeks. Endothelial dysfunction was evaluated using vasoreactivity assay, where acetylcholine-induced relaxation was assessed in phenylephrine pre-contracted rings. Combination therapy significantly improved vascular relaxation when compared with diabetic control as well as monotherapy. Restoration of nitrite levels along with prevention of collagen led to improved vasodilatation. Moreover, there was an overall reduction in aortic oxidative stress. We conclude that by simultaneously inhibiting ER stress and activating angiotensin-converting enzyme 2 deleterious effects of hyperglycemia on endothelium were significantly alleviated. This could serve as a novel strategy for the prevention of endothelial dysfunction.

The role of SARS-CoV-2 target ACE2 in cardiovascular diseases.

SARS-CoV-2, the virus responsible for the global coronavirus disease (COVID-19) pandemic, attacks multiple organs of the human body by binding to angiotensin-converting enzyme 2 (ACE2) to enter cells. More than 20 million people have already been infected by the virus. ACE2 is not only a functional receptor of COVID-19 but also an important endogenous antagonist of the renin-angiotensin system (RAS). A large number of studies have shown that ACE2 can reverse myocardial injury in various cardiovascular diseases (CVDs) as well as is exert anti-inflammatory, antioxidant, anti-apoptotic and anticardiomyocyte fibrosis effects by regulating transforming growth factor beta, mitogen-activated protein kinases, calcium ions in cells and other major pathways. The ACE2/angiotensin-(1-7)/Mas receptor axis plays a decisive role in the cardiovascular system to combat the negative effects of the ACE/angiotensin II/angiotensin II type 1 receptor axis. However, the underlying mechanism of ACE2 in cardiac protection remains unclear. Some approaches for enhancing ACE2 expression in CVDs have been suggested, which may provide targets for the development of novel clinical therapies. In this review, we aimed to identify and summarize the role of ACE2 in CVDs.

Reduced expression levels of heat shock protein 90 in a diminazene aceturate-resistant Babesia gibsoni isolate.

Heat shock protein 90 (HSP90) is a molecular chaperon and an essential component for stage differentiation and intracellular growth inside the host cells of many protozoans. HSP90 of Babesia gibsoni (BgHSP90) was suggested to function in the development of diminazene aceturate (DA)-resistance. Therefore, we examined the expression level of BgHSP90 in a DA-resistant B. gibsoni isolate. Transcription of the BgHSP90 gene in the DA-resistant isolate and wild-type B. gibsoni was assessed by quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). As a result, the copy number and relative amount of BgHSP90 transcripts in the DA-resistant isolate were significantly lower than those in the wild-type. Moreover, a rabbit anti-recombinant BgHSP90 antibody was developed, and the protein synthesis of BgHSP90 in the DA-resistant isolate was compared with that in the wild-type by Western blot analysis and indirect fluorescence assay. There was significantly less BgHSP90 protein than in the wild-type. Additionally, the relative intensity of BgHSP70 in DA-resistant isolate was also lower than that in the wild-type. This suggested that the expression of BgHSP90 and BgHSP70 in the DA-resistant B. gibsoni isolate was suppressed and that the reduced amount of BgHSP90 and BgHSP70 might cause the weak proliferation of the DA-resistant isolate. Further studies are necessary to elucidate the function of BgHSP90.

Diminazene Aceturate Stabilizes Atherosclerotic Plaque and Attenuates Hepatic Steatosis in apoE-Knockout Mice by Influencing Macrophages Polarization and Taurine Biosynthesis.

Atherosclerosis and nonalcoholic fatty liver disease are leading causes of morbidity and mortality in the Western countries. The renin-angiotensin system (RAS) with its two main opposing effectors, i.e., angiotensin II (Ang II) and Ang-(1-7), is widely recognized as a major regulator of cardiovascular function and body metabolic processes. Angiotensin-converting enzyme 2 (ACE2) by breaking-down Ang II forms Ang-(1-7) and thus favors Ang-(1-7) actions. Therefore, the aim of our study was to comprehensively evaluate the influence of prolonged treatment with ACE2 activator, diminazene aceturate (DIZE) on the development of atherosclerotic lesions and hepatic steatosis in apoE-/- mice fed a high-fat diet (HFD). We have shown that DIZE stabilized atherosclerotic lesions and attenuated hepatic steatosis in apoE-/- mice fed an HFD. Such effects were associated with decreased total macrophages content and increased α-smooth muscle actin levels in atherosclerotic plaques. Moreover, DIZE changed polarization of macrophages towards increased amount of anti-inflammatory M2 macrophages in the atherosclerotic lesions. Interestingly, the anti-steatotic action of DIZE in the liver was related to the elevated levels of HDL in the plasma, decreased levels of triglycerides, and increased biosynthesis and concentration of taurine in the liver of apoE-/- mice. However, exact molecular mechanisms of both anti-atherosclerotic and anti-steatotic actions of DIZE require further investigations.

Compounds from the Medicines for Malaria Venture Box Inhibit In Vitro Growth of Babesia divergens, a Blood-Borne Parasite of Veterinary and Zoonotic Importance.

Babesiosis is an infectious disease with an empty drug pipeline. A search inside chemical libraries for novel potent antibabesial candidates may help fill such an empty drug pipeline. A total of 400 compounds (200 drug-like and 200 probe-like) from the Malaria Box were evaluated in the current study against the in vitro growth of Babesia divergens (B. divergens), a parasite of veterinary and zoonotic importance. Novel and more effective anti-B. divergens drugs than the traditionally used ones were identified. Seven compounds (four drug-like and three probe-like) revealed a highly inhibitory effect against the in vitro growth of B. divergens, with IC50s ≤ 10 nanomolar. Among these hits, MMV006913 exhibited an IC50 value of 1 nM IC50 and the highest selectivity index of 32,000. The atom pair fingerprint (APfp) analysis revealed that MMV006913 and MMV019124 showed maximum structural similarity (MSS) with atovaquone and diminazene aceturate (DA), and with DA and imidocarb dipropionate (ID), respectively. MMV665807 and MMV665850 showed MMS with each other and with ID. Of note, a high concentration (0.75 IC50) of MMV006913 caused additive inhibition of B. divergens growth when combined with DA at 0.75 or 0.50 IC50. The Medicines for Malaria Venture box is a treasure trove of anti-B. divergens candidates according to the obtained results.

Acid exposure disrupts mucus secretion and impairs mucociliary transport in neonatal piglet airways.

Tenacious mucus produced by tracheal and bronchial submucosal glands is a defining feature of several airway diseases, including cystic fibrosis (CF). Airway acidification as a driving force of CF airway pathology has been controversial. Here we tested the hypothesis that transient airway acidification produces pathologic mucus and impairs mucociliary transport. We studied pigs challenged with intra-airway acid. Acid had a minimal effect on mucus properties under basal conditions. However, cholinergic stimulation in acid-challenged pigs revealed retention of mucin 5B (MUC5B) in the submucosal glands, decreased concentrations of MUC5B in the lung lavage fluid, and airway obstruction. To more closely mimic a CF-like environment, we also examined mucus secretion and transport following cholinergic stimulation under diminished bicarbonate and chloride transport conditions ex vivo. Under these conditions, airways from acid-challenged pigs displayed extensive mucus films and decreased mucociliary transport. Pretreatment with diminazene aceturate, a small molecule with ability to inhibit acid detection through blockade of the acid-sensing ion channel (ASIC) at the doses provided, did not prevent acid-induced pathologic mucus or transport defects but did mitigate airway obstruction. These findings suggest that transient airway acidification early in life has significant impacts on mucus secretion and transport properties. Furthermore, they highlight diminazene aceturate as an agent that might be beneficial in alleviating airway obstruction.

Therapeutic Effect of Diminazene Aceturate on Parasitic Blood Fluke Schistosoma mansoni Infection.

Praziquantel is currently the only drug available to treat schistosomiasis, a disease of enormous public health significance caused by a blood fluke of the genus Schistosoma Diminazene, a drug approved by the FDA, has been successfully used to treat diseases caused by blood protozoan parasites. In this study, we evaluated the antiparasitic properties of diminazene against Schistosoma mansoniex vivo and in mice harboring either chronic or early S. mansoni infections. In vitro, we monitored phenotypic and tegumental changes as well as the effects of the drug on pairing and egg production. In mice infected with either adult (chronic infection) or immature (early infection) worms, diminazene was administered intraperitoneally (10 to 100 mg/kg of body weight) or by oral gavage (100 to 400 mg/kg), and we studied the influence of the drug on worm burden and egg production. Liver and spleen pathologies and serum aminotransferase levels were also analyzed. In vitro, 50% effective concentrations (EC50) and EC90 revealed that diminazene is able to kill both immature and adult parasites, and its effect was time and concentration dependent. In addition, confocal laser scanning microscopy showed morphological alterations in the teguments of schistosomes. In an animal model, the influence of the drug on worm burden, egg production, hepatomegaly, and splenomegaly depended on the dosing regimen applied and the route of administration. Diminazene also caused a significant reduction in aminotransferase levels. Comparatively, diminazene treatment was more effective in chronic infection than in early infection. In tandem, our study revealed that diminazene possesses anthelmintic properties and inhibits liver injury caused by Schistosoma eggs.

ACE2 activation protects against cognitive decline and reduces amyloid pathology in the Tg2576 mouse model of Alzheimer's disease.

Mid-life hypertension and cerebrovascular dysfunction are associated with increased risk of later life dementia, including Alzheimer's disease (AD). The classical renin-angiotensin system (cRAS), a physiological regulator of blood pressure, functions independently within the brain and is overactive in AD. cRAS-targeting anti-hypertensive drugs are associated with reduced incidence of AD, delayed onset of cognitive decline, and reduced levels of Aß and tau in both animal models and human pathological studies. cRAS activity is moderated by a downstream regulatory RAS pathway (rRAS), which is underactive in AD and is strongly associated with pathological hallmarks in human AD, and cognitive decline in animal models of CNS disease. We now show that enhancement of brain ACE2 activity, a major effector of rRAS, by intraperitoneal administration of diminazene aceturate (DIZE), an established activator of ACE2, lowered hippocampal Aß and restored cognition in mid-aged (13-14-month-old) symptomatic Tg2576 mice. We confirmed that the protective effects of DIZE were directly mediated through ACE2 and were associated with reduced hippocampal soluble Aß42 and IL1-ß levels. DIZE restored hippocampal MasR levels in conjunction with increased NMDA NR2B and downstream ERK signalling expression in hippocampal synaptosomes from Tg2576 mice. Chronic (10 weeks) administration of DIZE to pre-symptomatic 9-10-month-old Tg2576 mice, and acute (10 days) treatment in cognitively impaired 12-13-month-old mice, prevented the development of cognitive impairment. Together these data demonstrate that ACE2 enhancement protects against and reverses amyloid-related hippocampal pathology and cognitive impairment in a preclinical model of AD.

ACE2 activator diminazene aceturate exerts renoprotective effects in gentamicin-induced acute renal injury in rats.

Acute Kidney Injury (AKI) comprises a rapidly developed renal failure and is associated with high mortality rates. The Renin-Angiotensin System (RAS) plays a pivotal role in AKI, as the over-active RAS axis exerts major deleterious effects in disease progression. In this sense, the conversion of Angiotensin II (Ang II) into Angiotensin-(1-7) (Ang-(1-7)) by the Angiotensin-converting enzyme 2 (ACE2) is of utmost importance to prevent worse clinical outcomes. Previous studies reported the beneficial effects of oral diminazene aceturate (DIZE) administration, an ACE2 activator, in renal diseases models. In the present study, we aimed to evaluate the therapeutic effects of DIZE administration in experimental AKI induced by gentamicin (GM) in rats. Our findings showed that treatment with DIZE improved renal function and tissue damage by increasing Ang-(1-7) and ACE2 activity, and reducing TNF-α. These results corroborate with a raising potential of ACE2 activation as a strategy for treating AKI.

The beneficial effects of angiotensin-converting enzyme II (ACE2) activator in pulmonary hypertension secondary to left ventricular dysfunction.

Pulmonary hypertension (PH) is a lethal and rapidly progressing disorder if left untreated, but there is still no definitive therapy. An imbalance between vasoconstriction and vasodilation has been proposed as the mechanism underlying PH. Among the vasomediators of the pulmonary circulation is the renin-angiotensin system (RAS), the involvement of which in the development of PH has been proposed. Within the RAS, angiotensin-converting enzyme 2 (ACE2), which converts angiotensin (Ang) II into Ang-(1-7), is an important regulator of blood pressure, and has been implicated in cardiovascular disease and PH. In this study, we investigated the effects of the ACE2 activator diminazene aceturate (DIZE) on the development of PH secondary to left ventricular dysfunction. A model of PH secondary to left ventricular dysfunction was established in 6-week-old Wistar rats by ascending aortic banding for 42 days. The hemodynamics and pulmonary expression of ACE, Ang II, ACE2, Ang-(1-7), and the Ang-(1-7) MAS receptor were investigated in the early treatment group, which was administered DIZE (15 mg/kg/day) from days 1 to 42, and in the late treatment group, administered DIZE (15 mg/kg/day) from days 29 to 42. Sham-operated rats served as controls. DIZE ameliorated mean pulmonary artery pressure, pulmonary arteriolar remodeling, and plasma brain natriuretic peptide levels, in addition to reversing the overexpression of ACE and up-regulation of both Ang-(1-7) and MAS, in the early and late treatment groups. DIZE has therapeutic potential for preventing the development of PH secondary to left ventricular dysfunction through ACEII activation and the positive feedback of ANG-(1-7) on the MAS receptor. A translational study in humans is needed to substantiate these findings.

Acute hypotensive effect of diminazene aceturate in spontaneously hypertensive rats: role of NO and Mas receptor.

Diminazene aceturate (DIZE) has been described as an angiotensin-converting enzyme 2 (ACE2) activator. We aimed to investigate DIZE effects on blood pressure (BP) of spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats. BP was recorded in awake and unrestrained rats 24 hours after femoral artery catheterization. DIZE (15 mg/kg, s.c.) produced a fast BP decrease only in SHR (P < .01). Pre-treatment with L-NAME (10 mg/kg, iv) did not change the hypotensive effect on systolic BP whereas mitigated the DIZE effect on diastolic BP (∆ Emax: -31 ± 5 DIZE vs -15 ± 1 mm Hg DIZE + L-NAME, P < .05). BP changes after DIZE remained unchanged after the treatment of rats with A-779 (50 ug/kg, iv), a Mas receptor blocker. Vasodilatation curves to DIZE (10-9 to 10-4  mol/L) in mesenteric arteries confirmed the NO-mediation on DIZE effects in SHR, as L-NAME (300 µmol/L) reduced the vascular sensitivity (∆EC50: -5.12 ± 0.09 CONTROL vs -4.66 ± 0.08 L-NAME, P < .05) and the magnitude of DIZE effect (area under the curve (AUC), 357.5 ± 8.2 DIZE vs 424.7 ± 11.6 L-NAME; P < .001), whereas A-779 (1 µmol/L) enhanced DIZE response (AUC, 357.5 ± 8.2 DIZE vs 309.8 ± 14.7 A-779, P < .05). Our findings indicate that DIZE acutely reduces the BP in SHR possibly by a mechanism other than Mas receptor activation. This effect seems to be mediated, at least partially, by NO.

Diminazene aceturate (DIZE) has cellular and in vivo antiarrhythmic effects.

Diminazene aceturate (DIZE) is an anti-protozoan compound that has been previously reported to increase the activity of the angiotensin-converting enzyme 2 (ACE2) and thus increase Angiotensin-(1-7) production, leading to cardioprotection against post-myocardial infarction dysfunction and structural remodelling. Moreover, DIZE is able to ameliorate morpho-functional changes after myocardial infarction by enhancing ACE2 activity, thus increasing Angiotensin-(1-7) production (a benefic peptide of the renin-angiotensin system). However, despite the improvement in cardiac function/structure, little is known about DIZE effects on arrhythmia suppression, contraction/excitable aspects of the heart and importantly its mechanisms of action. Thus, our aim was to test the acute effect of DIZE cardioprotection at the specific level of potential antiarrhythmic effects and modulation in excitation-contraction coupling. For this, we performed in vitro and in vivo techniques for arrhythmia induction followed by an acute administration of DIZE. For the first time, we described that DIZE can reduce arrhythmias which is explained by modulation of cardiomyocyte contraction and excitability. Such effects were independent of Mas receptor and nitric oxide release. Development of a new DIZE-based approach to ameliorate myocardial contractile and electrophysiological dysfunction requires further investigation; however, DIZE may provide the basis for a future beneficial therapy to post-myocardial infarction patients.

Occurrence of Trypanosoma spp. and piroplasm infections of horses at Obollo-Afor southeastern Nigeria and resistance profiles of trypanosomes to isometamidium and diminazene salts.

Epidemiological data on trypanosomosis and piroplasmosis of horses are lacking in southeastern Nigeria. The prevalence of trypanosome and piroplasm infections in horses and resistance profile of isolated trypanosomes to diminazene and isometamidium salts were investigated. For the cross-sectional study of horses billed for slaughter, 304 horses were randomly sampled. Approximately 2 ml of blood was collected into anticoagulant-treated bottles for haematocrit (HCT) determination, direct microscopic examinations, and rat inoculation. Gender, body condition scores (BCS), age groups, and body weights of sampled horses were noted. Two isolates of Trypanosoma brucei recovered from the cross-sectional study were profiled for resistance to isometamidium hydrochloride and diminazene diaceturate in 36 BALB/c mice. Standardized protocols were used (Eisler et al., Veterinary Parasitology 97:171-182, 2001). 19.1% of horses (95% confidence interval 14.7-23.5%) were positive for haemoparasite infections including Theileria equi (16.1%) and Babesia caballi (3.9%). Only two (0.66%) Trypanosoma brucei infections were seen, being from active cases. Associations between age or gender, and presence of haemoparasites were only random. Haemoparasite-infected horses had significantly (p < 0.05) lower mean HCT and body weights and poorer BCS. From resistance profiling, for each isolate, all mice in control groups were parasitaemic by day 6 post-inoculation, while mice in test groups remained aparasitaemic over 60-day observation period. The study showed the endemicity and weights of Trypanososma spp. and piroplasm infections and among horses within the area. Furthermore, circulating strains of Trypanosoma brucei in the area are still susceptible to isometamidium and diminazene salts in mice. The pharmacoepidemiological significances of these findings were discussed.

Spermine protects alpha-synuclein expressing dopaminergic neurons from manganese-induced degeneration.

Manganese exposure is among the many environmental risk factors linked to the progression of neurodegenerative diseases, such as manganese-induced parkinsonism. In animal models, chronic exposure to manganese causes loss of cell viability, neurodegeneration, and functional deficits. Polyamines, such as spermine, have been shown to rescue animals from age-induced neurodegeneration in an autophagy-dependent manner; nonetheless, it is not understood whether polyamines can prevent manganese-induced toxicity. In this study, we used two model systems, the Caenorhabditis elegans UA44 strain and SK-MEL-28 cells, both expressing the protein alpha-synuclein (α-syn) to determine whether spermine could ameliorate manganese-induced toxicity. Manganese caused a substantial reduction in the viability of SK-MEL-28 cells and hastened neurodegeneration in the UA44 strain. Spermine protected both the SK-MEL-28 cells and the UA44 strain from manganese-induced toxicity. Spermine also reduced the age-associated neurodegeneration observed in the UA44 strain compared with a control strain without α-syn expression and led to improved avoidance behavior in a functional assay. Treatment with berenil, an inhibitor of polyamine catabolism, which leads to increased intracellular polyamine levels, also showed similar cellular protection against manganese toxicity. While both translation blocker cycloheximide and autophagy blocker chloroquine caused a reduction in the cytoprotective effect of spermine, transcription blocker actinomycin D had no effect. This study provides new insights on the effect of spermine in preventing manganese-induced toxicity, which is most likely via translational regulation of several candidate genes, including those of autophagy. Thus, our results indicate that polyamines positively influence neuronal health, even when exposed to high levels of manganese and α-syn, and supplementing polyamines through diet might delay the onset of diseases involving degeneration of dopaminergic neurons.

Is the cardioprotective effect of the ACE2 activator diminazene aceturate more potent than the ACE inhibitor enalapril on acute myocardial infarction in rats?

Myocardial infarction is a major cause of cardiac dysfunction. All components of the cardiac renin-angiotensin system (RAS) are upregulated in myocardial infarction. Angiotensin-converting enzyme (ACE) and ACE2 are key enzymes involved in synthesis of components of RAS and provide a counter-regulatory mechanism within RAS. We compared the cardioprotective effect of the ACE2 activator diminazene aceturate (DIZE) versus the ACE inhibitor enalapril on post acute myocardial infarction (AMI) ventricular dysfunction in rats. Adult male rats received subcutaneous injections of either saline (control) or isoproterenol (85 mg/kg) to induce AMI. Rats with AMI confirmed biochemically and by ECG, were either left untreated (AMI) or administered DIZE (AMI + DIZE) or enalapril (AMI + enalapril) daily for 4 weeks. DIZE caused a significant activation of cardiac ACE2 compared with enalapril. DIZE caused a significantly greater enhancement of cardiac hemodynamics. DIZE also caused greater reductions in heart-type fatty acid binding protein (H-FABP), ß-myosin heavy chain (ß-MYH), and in heart mass to total body mass ratio. These results indicated that activation of cardiac ACE2 by DIZE enhanced the protective axis of RAS and improved myocardial function following AMI, whereas enalapril was not sufficient to restore all cardiac parameters back to normal.

Suspected resistance of Trypanosoma species to diminazene aceturate on a cattle farm in Nigeria.

African animal trypanosomiasis is a major cause of mortality and economic losses for the livestock industry in Nigeria. Chemotherapy has been the most reliable option for cattle herders, and the most commonly found drug on the market is diminazene aceturate. To ascertain the long-term efficacy of this compound, we sampled a cattle herd in Ogun State, Nigeria, 2 months after they were treated with diminazene aceturate. The ITS-PCR results revealed 19 positives for trypanosome DNA out of the 79 samples tested (24.1%, 95% CI 16.0-34.5). Seventeen out of the total 19 positives were Trypanosoma congolense (21.5%, 95% CI 13.9-31.8). Mixed infections were also observed. Therefore, the persistence of bovine trypanosomiasis at this Nigerian cattle farm despite treatment could be due to diminazene aceturate resistant trypanosomes being present in the herd.