Cytotoxicity and Autophagy Induced by Ivermectin via AMPK/mTOR Signaling Pathway in RAW264.7 Cells.

The widespread and excessive use of ivermectin (IVM) will not only cause serious environmental pollution, but will also affect metabolism of humans and other mammals that are exposed. IVM has the characteristics of being widely distributed and slowly metabolized, which will cause potential toxicity to the body. We focused on the metabolic pathway and mechanism of toxicity of IVM on RAW264.7 cells. Colony formation and LDH detection assay showed that IVM significantly inhibited the proliferation of and induced cytotoxicity in RAW264.7 cells. Intracellular biochemical analysis using Western blotting assay showed that LC3-B and Beclin-1 were upregulated and p62 was down-regulated. The combination of confocal fluorescence, calcein-AM/CoCl2, and fluorescence probe results showed that IVM could induce the opening of the mitochondrial membrane permeability transition pore, reduce mitochondrial content, and increase lysosome content. In addition, we focused on induction of IVM in the autophagy signal pathway. The Western blotting results showed that IVM increased expression of p-AMPK and decreased p-mTOR and p-S6K expression in protein levels, indicating that IVM activated the AMPK/mTOR signaling pathway. Therefore, IVM may inhibit cell proliferation by inducing cell cycle arrest and autophagy.

Conformational transitions and allosteric modulation in a heteromeric glycine receptor.

Glycine Receptors (GlyRs) provide inhibitory neuronal input in the spinal cord and brainstem, which is critical for muscle coordination and sensory perception. Synaptic GlyRs are a heteromeric assembly of α and ß subunits. Here we present cryo-EM structures of full-length zebrafish α1ßBGlyR in the presence of an antagonist (strychnine), agonist (glycine), or agonist with a positive allosteric modulator (glycine/ivermectin). Each structure shows a distinct pore conformation with varying degrees of asymmetry. Molecular dynamic simulations found the structures were in a closed (strychnine) and desensitized states (glycine and glycine/ivermectin). Ivermectin binds at all five interfaces, but in a distinct binding pose at the ß-α interface. Subunit-specific features were sufficient to solve structures without a fiduciary marker and to confirm the 4α:1ß stoichiometry recently observed. We also report features of the extracellular and intracellular domains. Together, our results show distinct compositional and conformational properties of α1ßGlyR and provide a framework for further study of this physiologically important channel.

Exploring drug repositioning for leishmaniasis treatment: Ivermectin plus polymeric micelles induce immunological response and protection against tegumentary leishmaniasis.

Leishmania amazonensis can cause a wide spectrum of the clinical manifestations of leishmaniasis in humans. The development of new therapeutics is a long and expensive task; in this context, drug repositioning could be considered a strategy to identify new biological actions of known products. In the present study, ivermectin (IVE) was tested against distinct Leishmania species able to cause disease in humans. In vitro experiments showed that IVE was effective to reduce the infection degree and parasite load in Leishmania donovani- and L. amazonensis-infected macrophages that were treated with it. In addition, using the culture supernatant of treated macrophages, higher production of IFN-γ and IL-12 and lower levels of IL-4 and IL-10 were found. Then, IVE was used in a pure form or incorporated into Poloxamer 407-based polymeric micelles (IVE/M) for the treatment of L. amazonensis-infected BALB/c mice. Animals (n = 16 per group) were infected and later received saline, empty micelles, amphotericin B (AmpB), IVE, or IVE/M. They were euthanized at one (n = 8 per group) and 30 (n = 8 per group) days after treatment and, in both endpoints, immunological, parasitological, and biochemical evaluations were performed. Results showed that both IVE and IVE/M induced higher levels of IFN-γ, IL-12, GM-CSF, nitrite, and IgG2a antibodies, as well as higher IFN-γ expression evaluated by RT-qPCR in spleen cell cultures. Such animals showed low organic toxicity, as well as significant reductions in the lesion's average diameter and parasite load in their infected tissue, spleen, liver, and draining lymph node. The efficacy was maintained 30 days post-therapy, while control mice developed a polarized Th2-type response and high parasite load. In this context, IVE could be considered as a new candidate to be applied in future studies for the treatment against distinct Leishmania species.

Ivermectin and moxidectin against soil-transmitted helminth infections.

Ivermectin and moxidectin, two macrocyclic lactones, are potent antiparasitic drugs currently registered and mainly used against filarial diseases; however, their potential value for improved soil-transmitted helminth (STH) control has been acknowledged. This review provides insights on recent studies evaluating the efficacy of ivermectin and moxidectin as single or coadministered therapy against human soil-transmitted helminthiases (including Strongyloides stercoralis infections) and on pharmacokinetic/pharmacodynamic parameters measured in treated populations. Furthermore, we discuss current gaps for research, highlight advantages - but also existing challenges - for uptake of ivermectin and/or moxidectin treatment schemes into routine STH control in endemic countries.

Structural Elucidation of Ivermectin Binding to α7nAChR and the Induced Channel Desensitization.

The α7 nicotinic acetylcholine receptor (α7nAChR) mediates signaling in the central nervous system and cholinergic anti-inflammatory pathways. Ivermectin is a positive allosteric modulator of a full-length α7nAChR and an agonist of the α7nAChR construct containing transmembrane (TMD) and intracellular (ICD) domains, but structural insights of the binding have not previously been determined. Here, combining nuclear magnetic resonance as a primary experimental tool with Rosetta comparative modeling and molecular dynamics simulations, we have revealed details of ivermectin binding to the α7nAChR TMD + ICD and corresponding structural changes in an ivermectin-induced desensitized state. Ivermectin binding was stabilized predominantly by hydrophobic interactions from interfacial residues between adjacent subunits near the extracellular end of the TMD, where the inter-subunit gap was substantially expanded in comparison to the apo structure. The ion-permeation pathway showed a profile distinctly different from the resting-state profile but similar to profiles of desensitized α7nAChR. The ICD also exhibited structural changes, including reorientation of the MX and h3 helices relative to the channel axis. The resulting structures of the α7nAChR TMD + ICD in complex with ivermectin provide opportunities for discovering new modulators of therapeutic potential and exploring the structural basis of cytoplasmic signaling under different α7nAChR functional states.

Enzyme-Responsive Lignin Nanocarriers for Triggered Delivery of Abamectin to Control Plant Root-Knot Nematodes (Meloidogyne incognita).

Intelligently responsive nanoparticles can improve insecticidal activity against target organisms and reduce the use of pesticides in agriculture. In this study, enzymatic hydrolysis lignin (EHL) nanocarriers with enzyme responsiveness were successfully prepared by electrostatic interaction, and abamectin (Abm)-loaded EHL-based nanoparticles (Abm@L-CL) were investigated. The release behavior of Abm@L-CL nanoparticles showed that Abm was released rapidly in the presence of cellulase and pectinase but slowly under natural conditions. The insecticidal activity of Abm@L-CL treatment (LC50 = 0.68 µg/mL) against nematodes (Meloidogyne incognita) was significantly more effective than that of original Abm treatment (LC50 = 1.32 µg/mL). The mortality rate of Abm@L-CL was more than 90% by applying the same dose of Abm after 12 h. The bioactivity of Abm@L-CL against root-knot nematodes was 1.7-fold greater than that of Abm. The result of fluorescence indicated that nanoparticles could enter the intestinal tract through the oral cavity of nematodes and achieve obvious gastric toxicity. Furthermore, the enzyme-controlled lignin-based Abm nanocarriers could penetrate the tomato root near the elongation zone. This study provided intelligent enzyme-responsive nanocarriers for efficient management of soil-borne diseases and pests in green agricultural inputs.

Synergism of macrocyclic lactones against Haemonchus contortus.

A possible synergistic effect of macrocyclic lactones' (MLs) combination has been previously described against resistant gastrointestinal nematodes of cattle. In addition to synergism, drug-drug interactions between MLs can also result in additive or antagonistic effect, considering the different MLs pharmacokinetics, pharmacodynamics, and interactions with molecular mechanisms of resistance. Therefore, the aim of the current work was evaluated the effect of different MLs combinations against Haemonchus contortus. Infecting larvae of two isolates (one susceptible and one resistant to ivermectin) were used in the larval migration inhibition test. After estimating the half maximal effective concentration of abamectin (ABA), eprinomectin, (EPR), ivermectin (IVM), and moxidectin (MOX) for both isolates, combinations were delineated by a simplex-centroid mixture experiment, and the mixture regression analysis was applied to the special cubic model. A synergistic effect was found for the EPR + MOX against the susceptible isolate as well as the EPR + MOX, IVM + MOX, and ABA + EPR + IVM against the resistant isolate. An antagonistic effect of ABA + IVM + MOX was found against the susceptible isolate. For the susceptible isolate, a higher inhibition was found with greater proportions of EPR and lower proportions of the other drugs compared to the reference mixture. For the resistant isolate, inhibition greater than that of the reference mixture was found with higher proportions of IVM as well as lower proportions of the other drugs. The synergistic and antagonistic effects were dependent on the following: (a) parasite drug resistance profile, (b) the composition of the combination, and (c) the proportions used, with EPR and IVM exerting a greater impact on these effects.

Anthelmintic resistance in sheep in the semiarid region of Minas Gerais, Brazil.

Anthelmintic efficacy was evaluated among sheep that had become naturally infected with gastrointestinal nematodes in 17 flocks located in the semiarid region of Minas Gerais, Brazil. Feces were collected individually from 1021 hairy sheep to determine the number of eggs per gram of feces (EPG) and for coprocultures to identify nematode genera the nematodes. Only the animals that presented EPG counts greater than or equal to 200 were included in the study (totaling 381 sheep). The animals were divided into three treatment groups: albendazole, ivermectin and levamisole. Fourteen days after the administration of anthelmintics, fecal samples were taken from all animals. In each flock, the pre-treatment and post-treatment arithmetic mean EPG were used to calculate the efficacy (FECR) for each of the treatment groups and the lower 95% confidence limit. Data were analyzed with the "eggCounts 2.3" package in RStudio, using a Bayesian model for paired design. The anthelmintics were classified as being efficacious (when the FECR was both equal to or above 95% and the lower 95% confidence limit was equal to or above 90%) or as encountering anthelmintic resistance (when the FECR was below 95% and the lower 95% confidence limit was below 90%) or inconclusive (when none of the other criteria were fulfilled). Albendazole and ivermectin were not effective in any of the flocks. Levamisole was effective against gastrointestinal nematodes in 25% of the flocks studied. Haemonchus, Trichostrongylus and Oesophagostomum genera were identified in this study in a semiarid region of Minas Gerais, Brazil. The genus Haemonchus was the most prevalent, followed by Trichostrongylus and Oesophagostomum. After anthelmintic treatment, the most prevalent genus was Haemonchus, followed by Trichostrongylus; the genus Oesophagostomum was not detected. The highest percentage of Haemonchus larvae was observed after treatment with ivermectin, followed by albendazole and levamisole. This study revealed the existence of gastrointestinal nematodes in sheep that present multiple resistance to all three main classes of anthelmintic drugs.

Effect of Ivermectin® on survivorship and fertility of Anopheles arabiensis in Ethiopia: an in vitro study.

BACKGROUND: Innovative vector control tools are needed to counteract insecticide resistance and residual malaria transmission. One of such innovative methods is an ivermectin (IVM) treatment to reduce vector survival. In this study, a laboratory experiment was conducted to investigate the effect of ivermectin on survivorship, fertility and egg hatchability rate of Anopheles arabiensis in Ethiopia. METHODS: An in vitro experiment was conducted using 3-5 days old An. arabiensis adults from a colony maintained at insectary of Tropical and Infectious Diseases Research Center, Jimma University (laboratory population) and Anopheles mosquitoes reared from larvae collected from natural mosquito breeding sites (wild population). The mosquitoes were allowed to feed on cattle blood treated with different doses of ivermectin (0 ng/ml, 5 ng/ml, 10 ng/ml, 20 ng/ml, 40 ng/ml and 80 ng/ml). During each feeding experiment, the mosquitoes were held in cages and blood-fed using a Hemotek feeder. Mortality and egg production were then recorded daily for up to 9 days. Time to death was analysed by a Cox frailty model with replicate as frailty term and source of mosquito (wild versus laboratory), treatment type (ivermectin vs control) and their interaction as categorical fixed effects. Kaplan Meier curves were plotted separately for wild and laboratory populations for a visual interpretation of mosquito survival as a function of treatment. RESULTS: Both mosquito source and treatment had a significant effect on survival (P < 0.001), but their interaction was not significant (P = 0.197). Compared to the controls, the death hazard of An. arabiensis that fed on ivermectin-treated blood was 2.3, 3.5, 6.5, 11.5 and 17.9 times that of the control for the 5 ng/ml, 10 ng/ml, 20 ng/ml, 40 ng/ml, and 80 ng/ml dose, respectively. With respect to the number of hatched larvae, hatched pupae and emerged adults per fed mosquitoes, a significant difference was found between the control and the 5 ng/ml dose group (P < 0.001). The number of hatched larvae and pupae, and emerged adults decreased further for the 10 ng/ml dose group and falls to zero for the higher doses. CONCLUSION: Treating cattle blood with ivermectin reduced mosquito survival, fertility, egg hatchability, larval development and adult emergence of An. arabiensis in all tested concentrations of ivermectin in both the wild and laboratory populations. Thus, ivermectin application in cattle could be used as a supplementary vector control method to tackle residual malaria transmission and ultimately achieve malaria elimination in Ethiopia.

Is the antiparasitic drug ivermectin a suitable candidate for the treatment of epilepsy?

There are only a few drugs that can seriously lay claim to the title of "wonder drug," and ivermectin, the world's first endectocide and forerunner of a completely new class of antiparasitic agents, is among them. Ivermectin, a mixture of two macrolytic lactone derivatives (avermectin B1a and B1b in a ratio of 80:20), exerts its highly potent antiparasitic effect by activating the glutamate-gated chloride channel, which is absent in vertebrate species. However, in mammals, ivermectin activates several other Cys-loop receptors, including the inhibitory γ-aminobutyric acid type A and glycine receptors and the excitatory nicotinic acetylcholine receptor of brain neurons. Based on these effects on vertebrate receptors, ivermectin has recently been proposed to constitute a multifaceted wonder drug for various novel neurological indications, including alcohol use disorders, motor neuron diseases, and epilepsy. This review critically discusses the preclinical and clinical evidence of antiseizure effects of ivermectin and provides several arguments supporting that ivermectin is not a suitable candidate drug for the treatment of epilepsy. First, ivermectin penetrates the mammalian brain poorly, so it does not exert any pharmacological effects via mammalian ligand-gated ion channels in the brain unless it is used at high, potentially toxic doses or the blood-brain barrier is functionally impaired. Second, ivermectin is not selective but activates numerous inhibitory and excitatory receptors. Third, the preclinical evidence for antiseizure effects of ivermectin is equivocal, and at least in part, median effective doses in seizure models are in the range of the median lethal dose. Fourth, the only robust clinical evidence of antiseizure effects stems from the treatment of patients with onchocerciasis, in which the reduction of seizures is due to a reduction in microfilaria densities but not a direct antiseizure effect of ivermectin. We hope that this critical analysis of available data will avert the unjustified hype associated with the recent use of ivermectin to control COVID-19 from recurring in neurological diseases such as epilepsy.

EVALUATION OF INTRACARDIAC ADMINISTRATION OF POTASSIUM CHLORIDE, IVERMECTIN, OR LIDOCAINE HYDROCHLORIDE FOR EUTHANASIA OF ANESTHETIZED BLUE CRABS (CALLINECTES SAPIDUS).

Methods to anesthetize and euthanize aquatic invertebrates have proven unreliable in decapods; thus studies to optimize euthanasia techniques for crustaceans are needed. Study objectives were to evaluate efficacy of intracardiac potassium chloride (KCl), ivermectin, or lidocaine hydrochloride (HCl) for euthanasia of anesthetized blue crabs (Callinectes sapidus). Twenty adult male crabs (n = 5/group) were immersed in 500 mg/L eugenol for 5 min beyond loss of the righting reflex and then randomly administered intracardiac 10 mEq/kg KCl (333 mg/mL), 5 mg/kg ivermectin (10 mg/ml), 100 mg/kg lidocaine hydrochloride (HCl) (20 mg/ml), or 5 ml/kg saline (0.9%). Serial heart rate assessments were made using a Doppler probe placed over the dorsum, and times to loss of righting reflex, Doppler sound cessation, and/or recovery were recorded. Median (range) time to loss of righting reflex was 32 (17-57) min. One crab in all groups, except lidocaine HCl, had no detectable Doppler sounds prior to injection. In the remaining crabs, Doppler sound cessation occurred in 4/4, 4/4, 4/5, and 0/4 crabs administered KCl, ivermectin, lidocaine HCl, and saline, respectively. Median (range) time to Doppler sound cessation was 30 (0-55), 18 (16-28), and 50 (0-90) s in KCl, ivermectin, and lidocaine HCl groups, respectively. Tonic limb movements were observed in 5/5 KCl-treated crabs. Median (range) time to recovery was 180 (115-345) and 300 m in four saline-treated crabs and one lidocaine HCl-treated crab, respectively. Intracardiac KCl at 10 mEq/kg and ivermectin at 5 mg/kg were effective, rapid methods for euthanasia of anesthetized blue crabs.

Effect of cyclosporin A on the toxicity of ivermectin, eprinomectin and moxidectin in populations of Rhipicephalus microplus.

Rhipicephalus microplus is mainly controlled by acaricides. However, reports of resistance to acaricides including macrocyclic lactones (MLs) have become frequent worldwide. Involvement of ABC transporters (ABCts) in populations resistant to ivermectin has been demonstrated. Thus, the aim of this study was to evaluate the efficacy of ivermectin, eprinomectin and moxidectin with and without use of synergistic cyclosporin A (CsA) in resistant populations of R. microplus using larval immersion tests (LITs). Engorged females were collected from four farms in the semiarid region of northeastern Brazil that had histories of continuous use of ivermectin. Questionnaires were applied to collect information about management aimed at controlling ticks on these farms. Resistance to MLs was observed on all of the farms. There was statistically significant synergism (p < 0.05) between CsA and ivermectin in all populations; between CsA and eprinomectin in only one population; and between CsA and moxidectin in two populations. It was concluded that, despite the involvement of ABCts in the mechanisms of resistance to ivermectin, metabolic detoxification does not seem to be the mechanism predominantly involved in resistance to eprinomectin and moxidectin in the populations of R. microplus evaluated.

Clinical and Parasitological Evaluation of Ivermectin and Ivermectin + Pyrantel Against Oxyuris Equi in Equines.

The equine pinworm could become an increasingly common problem, as there are reports of failure in the control of this parasite. The aim of this study was to evaluate the effects of ivermectin (IVM) and IVM combined with pyrantel pamoate (PYR). Thirteen parasitological positive equines were treated with oral IVM (200 µg/kg) and therapeutic efficacy, clinical recovery and the egg reappearance period (ERP) were evaluated. In cases for which ERP was shorter than the pre-patent period (PPP), a second treatment was performed with IVM (200 µg/kg) + PYR (6.6 mg/kg), followed by the same evaluation criteria described above. Therapeutic efficacy was 100% with IVM + PYR and 53.84% with IVM. The mean ERP was shorter than the PPP with both formulations, 77.55 days with IVM + PYR and 50 days with IVM. The presence of egg mass was always associated with a least one clinical sign. The reduction in the number of clinical signs per animal from Day 0 to Day 30 was greater in equines treated with IVM + PYR compared to those treated with IVM alone. The animals treated with IVM were 4.5-fold more likely to present clinical signs 30 days after treatment than those treated with IVM+PYR. A negative correlation was found between ERP and the number of clinical signs at 30 days in the animals treated with IVM. This clinical and parasitological evaluation demonstrated that the combination of IVM+PYR was more effective than IVM alone to control Oxyuris equi.

Ivermectin Attenuates CCl4-Induced Liver Fibrosis in Mice by Suppressing Hepatic Stellate Cell Activation.

Liver fibrosis, a common liver dysfunction with high morbidity and mortality rates, is the leading cause of cirrhosis and hepatocellular carcinoma, for which there are no effective therapies. Ivermectin is an antiparasitic drug that also has been showing therapeutic actions in many other diseases, including antiviral and anticancer actions, as well as treating metabolic diseases. Herein, we evaluated the function of ivermectin in regulating liver fibrosis. Firstly, carbon tetrachloride (CCl4)-injected Balb/c mice were used to assess the antifibrosis effects of ivermectin in vivo. Further, CFSC, a rat hepatic stellate cell (HSC) line, was used to explore the function of ivermectin in HSC activation in vitro. The in vivo data showed that ivermectin administration alleviated histopathological changes, improved liver function, reduced collagen deposition, and downregulated the expression of profibrotic genes. Mechanistically, the ivermectin treatment inhibited intrahepatic macrophage accumulation and suppressed the production of proinflammatory factors. Importantly, the ivermectin administration significantly decreased the protein levels of α-smooth muscle actin (α-SMA) both in vivo and in vitro, suggesting that the antifibrotic effects of ivermectin are mainly due to the promotion of HSC deactivation. The present study demonstrates that ivermectin may be a potential therapeutic agent for the prevention of hepatic fibrosis.

Evaluation of a treatment protocol in dogs with intraspinal spirocercosis.

OBJECTIVE: To evaluate the efficiency and safety of a doramectin-based treatment protocol in dogs affected by intraspinal spirocercosis (Spirocerca lupi). ANIMALS: Client-owned dogs that were admitted to a veterinary hospital during 2021 to 2022 with acute onset of neurological signs and diagnosed with intraspinal spirocercosis. All dogs underwent complete neurological evaluation, CSF analysis, PCR confirmation of CNS S lupi infection, and follow-up evaluation of at least 6 months. PROCEDURES: Upon diagnosis, dogs were treated with doramectin at a dose of 400 µg/kg, SC, q 24 h for 3 consecutive days, followed by the same dose once a week for 6 weeks. Prednisone was administered at a dose of 1 mg/kg, PO, q 24 h and tapered every 3 days. Antimicrobial clindamycin was administered at a dose of 12.5 mg/kg, PO, q 12 h for 7 days to reduce the risk of secondary spinal cord infection. Short- and long-term outcomes (1 week to 56 months) were recorded. RESULTS: 8 dogs fulfilled the inclusion criteria, 7 of which presented with neurological deficits and 1 with cervical pain. Initiation of treatment was associated with stopping the deterioration in 7 of 8 dogs. Seven dogs improved and 6 recovered ambulation. One dog was euthanized due to lack of improvement. Six of the recovered dogs were still ataxic on the last follow-up examination at 6 to 56 months. No adverse effects of the drug were noted. CLINICAL RELEVANCE: Frequent administration of doramectin was found to be safe and effective in preventing neurological deterioration in dogs with intraspinal spirocercosis.

Repurposing Ivermectin to augment chemotherapy's efficacy in osteosarcoma.

BACKGROUND: Osteosarcoma is the most frequent malignant bone malignancy and the current treatments are ineffective. Ivermectin, an anti-protozoal drug, has been shown to have anti-cancer activity. This work investigated the potential of repurposing ivermectin to augment chemotherapy's efficacy in osteosarcoma. METHODS: Proliferation, migration and apoptosis assays were performed in ivermectin-treated osteosarcoma cells. Combination studies were performed. Osteosarcoma xenograft mouse model was established to investigate the in vivo efficacy of ivermectin. Intracellular reactive oxygen species (ROS) and mitochondrial superoxide, membrane potential, ATP, 8-OHdG level, protein carbonylation and lipid peroxidation were determined after ivermectin treatment. RESULTS: Ivermectin was effective and acted synergistically with doxorubicin in osteosarcoma cells regardless of cellular origin and genetic profiling. This was achieved through suppressing inhibiting growth and migration, and inducing caspase-dependent apoptosis. Ivermectin also significantly inhibited osteosarcoma growth in vivo and its combination with doxorubicin resulted in much greater efficacy than doxorubicin alone. Importantly, the effective dose of ivermectin was clinically feasible and did not cause significant toxicity in mice. Mechanistical analysis showed that ivermectin induced oxidative stress and damage, and mitochondrial dysfunction. CONCLUSIONS: Our findings indicate that ivermectin has utility in treating patients with osteosarcoma, especially those resistant to chemotherapy.

Ivermectin Attenuates CCl4-Induced Liver Fibrosis in Mice by Suppressing Hepatic Stellate Cell Activation.

Liver fibrosis, a common liver dysfunction with high morbidity and mortality rates, is the leading cause of cirrhosis and hepatocellular carcinoma, for which there are no effective therapies. Ivermectin is an antiparasitic drug that also has been showing therapeutic actions in many other diseases, including antiviral and anticancer actions, as well as treating metabolic diseases. Herein, we evaluated the function of ivermectin in regulating liver fibrosis. Firstly, carbon tetrachloride (CCl4)-injected Balb/c mice were used to assess the antifibrosis effects of ivermectin in vivo. Further, CFSC, a rat hepatic stellate cell (HSC) line, was used to explore the function of ivermectin in HSC activation in vitro. The in vivo data showed that ivermectin administration alleviated histopathological changes, improved liver function, reduced collagen deposition, and downregulated the expression of profibrotic genes. Mechanistically, the ivermectin treatment inhibited intrahepatic macrophage accumulation and suppressed the production of proinflammatory factors. Importantly, the ivermectin administration significantly decreased the protein levels of α-smooth muscle actin (α-SMA) both in vivo and in vitro, suggesting that the antifibrotic effects of ivermectin are mainly due to the promotion of HSC deactivation. The present study demonstrates that ivermectin may be a potential therapeutic agent for the prevention of hepatic fibrosis.

Modulation of P2X4 receptor activity by ivermectin and 5-BDBD has no effect on the development of ARPKD in PCK rats.

Autosomal recessive polycystic kidney disease (ARPKD) is an inherited pathology caused mainly by mutations of the polycystic kidney and hepatic disease 1 (PKHD1) gene, which usually leads to end-stage renal disease. Previous studies suggested that the P2X purinoreceptor 4 (P2X4 R) may play an important role in the progression of ARPKD. To test this hypothesis, we assessed the chronic effects of ivermectin (P2X4 R allosteric modulator) and 5-BDBD (P2X4 R antagonist) on the development of ARPKD in PCK/CrljCrl-Pkhd1pck/CRL (PCK) rats. Our data indicated that activation of ATP-mediated P2X4 R signaling with ivermectin for 6 weeks in high dose (50 mg/L; water supplementation) decreased the total body weight of PCK rats while the heart and kidney weight remained unaffected. Smaller doses of ivermectin (0.5 or 5 mg/L, 6 weeks) or the inhibition of P2X4 R signaling with 5-BDBD (18 mg/kg/day, food supplement for 8 weeks) showed no effect on electrolyte balance or the basic physiological parameters. Furthermore, cystic index analysis for kidneys and liver revealed no effect of smaller doses of ivermectin (0.5 or 5 mg/L) and 5-BDBD on the cyst development of PCK rats. We observed a slight increase in the cystic liver index on high ivermectin dose, possibly due to the cytotoxicity of the drug. In conclusion, this study revealed that pharmacological modulation of P2X4 R by ivermectin or 5-BDBD does not affect the development of ARPKD in PCK rats, which may provide insights for future studies on investigating the therapeutic potential of adenosine triphosphate (ATP)-P2 signaling in PKD diseases.

Signs of multiple anthelmintic resistance in sheep gastrointestinal nematodes in Sweden.

Gastrointestinal nematodes in small ruminants are clinically and economically important parasites that often are controlled with anthelmintics. In this study, we compiled information on the anthelmintic efficacy collected on sheep farms according to routines established by Farm & Animal Health in Sweden. The efficacies of benzimidazoles (i.e. albendazole or fenbendazole, n = 30), ivermectin (n = 47), levamisole (n = 2) or moxidectin (n = 2) were examined between 2015 and 2021 in 81 treatment groups on 49 non-randomly selected farms in south-central Sweden. Drug efficacies were estimated with the faecal egg count reduction test. In addition, efficacy data were in most cases supplemented with data on the abundance of the three most common nematode genera in sheep by performing droplet digital (dd) PCR on coprocultures. Efficacies of <95% for benzimidazoles or ivermectin were identified in 37% and 77% of the tested groups, respectively. In addition, on 27 (55%) of the 49 farms where both benzimidazoles and ivermectin were tested, multiple resistance was found on 8 (30%). In contrast, on each of the two farms tested for levamisole and moxidectin both drugs proved to be 100% effective. However, because post-sampling was performed earlier than recommended in several susceptible groups (benzimidazoles = 15, and ivermectin = 10 groups), this could have underestimated the severity of the situation. Mainly larvae from the genus Haemonchus were detected in post-treatment coprocultures, in all groups with declared resistance, suggesting that this parasite was primarily associated with anthelmintic resistance. Unexpectedly, the DNA of larvae, which survived treatment, was also detected on farms declared as susceptible. Taken together, this indicates that the situation regarding the anthelmintic efficacy has deteriorated compared with the latest nationwide study on Swedish sheep farms conducted more than a decade ago. Unlike the previous study, the farm selection here was not strictly randomized but rather opportunistic i.e., only farms with a recognized parasite problem were included. Thus, there is a need for a truly randomized study to get an update on the extent of the situation of anthelmintic resistance at a national level, as well as to identify risk factors involved in the resistance selection. Research is also required to establish the optimal intervals for sampling post-treatment.

Conservation of Importin α Function in Apicomplexans: Ivermectin and GW5074 Target Plasmodium falciparum Importin α and Inhibit Parasite Growth in Culture.

Signal-dependent transport into and out of the nucleus mediated by members of the importin (IMP) superfamily of nuclear transporters is critical to the eukaryotic function and a point of therapeutic intervention with the potential to limit disease progression and pathogenic outcomes. Although the apicomplexan parasites Plasmodium falciparum and Toxoplasma gondii both retain unique IMPα genes that are essential, a detailed analysis of their properties has not been performed. As a first step to validate apicomplexan IMPα as a target, we set out to compare the properties of P. falciparum and T. gondii IMPα (PfIMPα and TgIMPα, respectively) to those of mammalian IMPα, as exemplified by Mus musculus IMPα (MmIMPα). Close similarities were evident, with all three showing high-affinity binding to modular nuclear localisation signals (NLSs) from apicomplexans as well as Simian virus SV40 large tumour antigen (T-ag). PfIMPα and TgIMPα were also capable of binding to mammalian IMPß1 (MmIMPß1) with high affinity; strikingly, NLS binding by PfIMPα and TgIMPα could be inhibited by the mammalian IMPα targeting small molecules ivermectin and GW5074 through direct binding to PfIMPα and TgIMPα to perturb the α-helical structure. Importantly, GW5074 could be shown for the first time to resemble ivermectin in being able to limit growth of P. falciparum. The results confirm apicomplexan IMPα as a viable target for the development of therapeutics, with agents targeting it worthy of further consideration as an antimalarial.