Evaluation of mechanical transmission of Trypanosoma vivax by Stomoxys calcitrans in a region without a cyclic vector.

This work investigated the mechanical transmission of Trypanosoma vivax by Stomoxys calcitrans to cattle in a region without a cyclic vector. The study involved two experiments, one with calves experimentally infected with T. vivax, in the acute phase of trypanosomosis (Experiment 1) and the other in the chronic phase (Experiment 2). In both experiments, two transmission methods were used with flies that had not fed for 24 h or had never fed: (i) Method 1: flies released freely in cattle pens (≈3,300 flies/pen for 10 days); and (ii) Method 2: flies placed in a feeding chamber (12 flies/animal). To develop Method 1 in the two experiments (acute and chronic phases), T. vivax-positive animals were kept with T. vivax-negative animals. Periodically, the Brener method, Woo method, blood smears, cPCR, ELISA, IFAT, and Imunoteste® were performed to detect T. vivax in the animals. We also recorded the animals' head tossing and hoof stomping and the number of flies near the pens' inner walls. Subsequently, biological testing was performed using lambs. For Method 2 in both experiments, flies inside the feeding chamber first fed on T. vivax-positive animals and later on negative animals. In both experiments and methods, we examined the flies for the presence of T. vivax through blood smears and cPCR of the proboscis and abdomen. In Experiment 2 (chronic phase), a test was conducted to determine how long trypomastigotes forms could survive on the blood of animals with different levels of parasitemia. None of the animals (calves and lambs) became infected with T. vivax or showed antibodies against it. During the evaluation period, the animals in the presence of the flies exhibited more hoof stomping and head tossing compared to those without flies (control). Additionally, there was an increase in the number of flies in the pens during the experiment. Only in Experiment 1 (acute phase) were T. vivax trypomastigotes and DNA found in the abdomen of the flies but not in the proboscis. In Experiment 2 (chronic phase), higher concentrations of trypomastigotes per milliliter of blood were associated with a shorter the lifespan of this stage of the parasite. In conclusion, under the variable conditions of the experiments (hosts, number of flies, and level of parasitemia), S. calcitrans was unable to mechanically transmit T. vivax to cattle.

What is the optimal timing to initiate strategic control of Rhipicephalus microplus in taurine cattle in a tropical region?

Cattle tick control poses a significant challenge for livestock in tropical and subtropical regions. The objective of this study was to determine the most suitable timing to initiate a strategic tick control program and to identify the ideal number of acaricide treatments for adult taurine cattle (Bos taurus taurus) in a tropical region throughout the year. Three groups with 10 bovines each were performed: T01 (strategic treatment in late autumn/winter/late spring, every 28 days), T02 (strategic treatment to act in the "first tick generation" - early spring/summer/early autumn, every 28 days) and T03 (control). Tick counts (females 4-8 mm) were conducted every 14 days. If the tick burden in any group reached 30 or more during these counts, we applied an additional treatment. Over the course of a year, T02 required significantly fewer (p < 0.05) acaricide treatments than T01, with nine treatments for T02 and eleven for T01. Furthermore, during the tick counts, animals in T02 showed a lower tick burden compared to those in T01. Initiating the strategic tick control program in early spring, corresponding to the first tick generation, proved more effective than starting in autumn. This approach not only required fewer acaricide treatments but also resulted in a reduced tick burden. These benefits are particularly valuable in terms of animal welfare and managing acaricide resistance issues.

Anaplasmosis outbreaks in taurine and zebu cattle in Brazil: causes, treatments, and Anaplasma marginale genetic diversity.

This study aimed to investigate seven outbreaks of A. marginale infection in two regions of Brazil, affecting taurine, zebu, and crossbred cattle. We assessed the possible causes, treatment measures, and genetic diversity of A. marginale. These outbreaks occurred in two states (Goiás: outbreaks 1-7; Mato Grosso do Sul: outbreak 3), breeds (Holstein, Nellore, and crossbreed), age groups (beef cattle: 18-25 days old and 7-8 months; dairy cattle: 18-25 days old, 13-14 months, and cow after the first birth) and rearing systems (feedlot, pasture, pen in a wood shaving bedding system and compost bedded-pack barns). Metaphylactic or prophylactic treatments varied according to outbreak (imidocarb dipropionate: outbreaks 1-4 and 6; enrofloxacin: outbreaks 5 and 7; diminazene diaceturate: outbreak 5). In outbreaks 6 and 7, the packed cell volume was monitored. In all outbreaks, the practice of needle/syringe sharing was discontinued. For outbreaks 1-3, clinical signs and mortality (range, 4.8-13.3%) occurred 36-45 days after entry into the feedlot. In outbreak 4, A. marginale was diagnosed in 66.2% of the calves (bacteremia, 0-4.5%), with a mortality of 8.6%. Among nursing calves aged 60 days during outbreak 5, 53.8% were infected with A. marginale, with average bacteremia of 2.7% (range, 0-21.3%), and a mortality of 13.8%. In dairy heifers aged 14 months, raised in paddocks lacking vegetation cover and infested with R. microplus, then transitioned to a rotational grazing system also infested with R. microplus, the A. marginale bacteremia ranged from 3.2 to 6.7%, with a mortality of 20%. Before monitoring during outbreak 7, the mortality was 17.9%, but no further deaths were observed after monitoring initiation. In conclusion, possible causes triggering the outbreaks included primary tick infestation, needle/syringe sharing, and stress factors which may have affected the immunological statues of animals in the feedlots. Control measures performed in all outbreaks were effective. The partial msp4 gene sequences of A. marginale generated herein belonged to two haplotypes, but further research would be needed to investigate if this finding has any clinical significance.

Influence of the acaricide emulsion pH on the effectiveness of spray products to control the cattle tick: laboratory and field investigations.

The current work evaluated the efficacy of 10 commercial acaricides in different pHs (4.5, 5.5, and 6.5) in laboratory (adult immersion tests (AIT), pH evaluation over time) and field assays (tick counts and efficacy). In the AIT (n=70), higher efficacies were obtained when the acaricide emulsion had a more acidic pH (4.5), mainly for two combinations of pyrethroids + organophosphate (acaricide 3 and acaricide 9). For amidine, a higher pH (6.5) showed a higher efficacy. Over time, there was a trend in the pH of these emulsions increasing. When the efficacy of chlorpyrifos + cypermethrin + piperonyl butoxide (acaricide 3) at different pHs was evaluated over time (0, 6, 12, and 24h) by AIT, the less acidic pH (6.5) showed a strongly variation in the acaricide efficacy range. The mean pH of the water samples from different regions of Brazil was 6.5. In the field, the association of pyrethroid + organophosphates (acaricide 9) with pH of 4.5 and 5.5 were more effective in tick control than the emulsion prepared with this same spray formulation at pH 6.5. The pH of the acaricide emulsions is an important point of attention and is recommended that the veterinary industry start to develop/share information regarding how the pH can affect the acaricide efficacy.

Effect of a preventive strategic control program, with imidocarb dipropionate, against tick fever agents in dairy calves.

Chemoprophylaxis with dipropionate imidocarb (IMD) is a method adopted to prevent cattle tick fever (TF). Sixty weaned dairy heifers (±60 days old), without previous exposure to Rhipicephalus microplus ticks, were housed in Tifton paddocks and were subsequently exposed to R. microplus ticks and monitored up to 315 days old. Thirty animals were kept as controls (T01) and 30 received five preventive strategic treatments with IMD at 21-day intervals (T02). The heifers were monitored weekly by means of packed cell volume (PCV) and blood smears to evaluate the presence of TF agents. Salvage treatments (ST) with diminazene and enrofloxacin were administered when animals showed PCV ≤ 24%. The A. marginale prevalence was 39.3% and 37.7%, B. bovis 6.0%, and 7.3%, and B. bigemina 16.3% and 13.7% for T01 and T02, respectively. Regarding PCV values, group T01 showed lower PCV than group T02, between 119 and 161 days of life, but when animals were 196, 210, 217, and between 252 to 301 days old, an inversion occurred. The IMD treatment protocol was effective in group T02 from day 91 to 175 while treatment was being administered, but from day 182 to 315 after the IMD treatment protocol was completed, the number of salvage treatments against TF agents performed in T02 group increased significantly. The sequential application of IMD treatments with intervals less than 21 days is not recommended.

How many cattle can be infected by Trypanosoma vivax by reusing the same needle and syringe, and what is the viability time of this protozoan in injectable veterinary products?

It was investigated how many cattle become infected with Trypanosoma vivax by subcutaneous (SC), intramuscular (IM) and intravenous (IV) routes, using the same syringe and needle from an animal with acute T. vivax infection. Besides, the T. vivax viability in 109 injectable veterinary drugs (antibiotics, antiparasitics, reproductive hormones, vitamin complex and derivatives, vaccines, anaesthetics, anti-inflammatory/antipyretics, antitoxics). In the field assay, four groups were performed: T01, T02 and T03 animals that received saline solution with the same syringe and needle contaminated with T. vivax via SC, IM and IV routes, respectively, and T04 control animals that received only saline solution with the same syringe and needle IV. In the laboratory, drugs had their pH measured and T. vivax viability verified. The number of cattle infected with T. vivax via SC (3/20) was lower (P ≤ 0.05) compared to via IM (9/20), which was lower (P ≤ 0.05) compared to IV (15/20). The solution pH did not influence T. vivax viability. In 44% (48/109) of the products, T. vivax remained viable regardless of time, stooding out that in 100% of oxytocins the protozoan was verified, at some evaluation times. The mean of T. vivax quantified in foot-and-mouth and brucellosis vaccines and in doramectin-based products were higher (P ≤ 0.05) than found in blood + saline solution.

Long-term efficacy of toltrazuril in naïve calves prophylactically treated and experimentally infected with Eimeria spp.

Eimeria are ubiquitous parasites and eimeriosis treatment is based on coccidiostats or coccidicides used prophylactically, metaphylactically, or therapeutically. The long-term efficacy of toltrazuril (TZR, 15 mg/kg) against experimentally infected naïve calves was investigated. Seven groups (six treated and one control) of six animals each were formed. Animals of each group received a single TZR prophylactic oral treatment on days - 42, - 35, - 28, - 21, - 14, and - 7 before the challenge with infectious sporulated oocysts of Eimeria spp. (100,000 oocysts: 59.5% E. zuernii, 38.1% E. bovis, 1.2%, E. alabamensis, and 1.2% E. ellipsoidalis). The long-term efficacy was assessed based on Eimeria spp. oocyst excretion by fecal oocyst counts. Three calves from the control group presented diarrhea with blood, which was not observed in animals belonging to the treatment groups. The TZR achieved efficacy greater than 95.0% up to 14 and 7 days. This formulation showed efficacy above 95% for 7 to 14 days, between 82 and 84% for 21 to 28 days and between 50 and 64% for 35 to 42 days.

Scanning electron microscopic evaluation of microgap size in Morse taper implants with straight and angled abutments.

The aim of this study was to compare the sizes of the implant-abutment microgap in Morse taper implants with a straight abutment (ISA group) and Morse taper implants with an angled abutment (IAA group). A total of 19 Morse taper dental implants (3.8 × 11.0 mm) were used with their respective abutments; 10 abutments were angled at 20°, and 9 were straight. The implant-abutment units were immersed in an epoxy resin to form a base and then cross-sectioned until about 50% of the volume was removed. The specimens were analyzed under a scanning electron microscope, and the size of the microgap was measured at 3 regions on each side of the implant, for a total of 6 measurements per specimen. Mann-Whitney U and Kruskal-Wallis tests were used for statistical analysis. In the ISA group, there were no statistically significant differences in the median sizes of the microgap at the different regions of the interface (P > 0.05; Kruskal-Wallis test). The mean (SD) values of the largest and smallest microgaps in the ISA group were 1.64 (1.38) and 0.83 (0.41) µm, respectively. In the IAA group, there was a statistically significant difference in the median sizes of the microgaps at the upper and middle regions on the left side (P = 0.031; Kruskal-Wallis test). The mean (SD) values of the largest and smallest micrograps were 1.43 (0.062) and 0.61 (0.27) µm, respectively. Comparison of the different regions in the ISA and IAA groups revealed that the only statistically significant difference was at the lower region on the right side (P = 0.027; Mann-Whitney U test). The sizes of the microgaps in the ISA and IAA groups were statistically similar in an overall analysis (P > 0.05; Mann-Whitney U test); however, the IAA group showed greater sealing ability. In addition, in both groups in vitro titanium oxide formation was observed at the interface, characterizing a type of cold-weld joint that provides effective implant-abutment sealing.

An update on amitraz efficacy against Rhipicephalus microplus after 15 years of disuse.

Amitraz is an acaricide that is widely used in veterinary medicine to control the cattle tick Rhipicephalus microplus. However, controversy exists in the literature regarding the resistance of R. microplus to this product. The present work provides an update on the acaricidal efficacy of amitraz (Triatox®, 12.5 % amitraz) after 15 years without its use on a property. Two in vivo (bovines treated with amitraz and submitted to tick counts, n = 20 animals) and one in vitro (adult immersion test, n = 40 ticks) assays were performed to determine product efficacy. The efficacy of the commercial formulation tested in the first in vivo trial ranged from 14.1 to 47.0%, and in the second from 3.6 to 35.1%, for the 28 days of the experiments. Efficacy for the in vitro trial was 47.38%. The dose recommended by the manufacturer of the product did not cause mortality to most of the ticks of this strain, and efficacy/resistance was not reverted or modified after 15 years (estimated 60 tick generations).

Delayed reduction of Anaplasma marginale parasitemia and packed cell volume normalization despite prolonged enrofloxacin treatment of cattle co-infected with Trypanosoma vivax.

Although co-infections of Trypanosoma vivax, Anaplasma spp., and Babesia spp. have been reported, knowledge gaps remain that need to be addressed. The present study evaluated the efficacy of enrofloxacin (7.5 mg/kg) against A. marginale in naturally infected cattle and cattle experimentally co-infected with T. vivax by observation of the variation in A. marginale parasitemia and packed cell volume (PCV) for 39 days. Bovines were distributed into two groups, each with six calves: T01 = animals immunosuppressed with dexamethasone and with latent anaplasmosis; T02 = animals immunosuppressed with dexamethasone, with latent anaplasmosis and experimentally co-infected with T. vivax on day 0 (D0). Animals of both groups were immunosuppressed with dexamethasone and received enrofloxacin (7.5 mg/kg) whenever mean values of parasitemia for A. marginale were ≥ 5% per group. Cattle of group T02 were also treated with isometamidium chloride (0.5 mg/kg) on D25. On D17 and D22 to D28 of the study, there was a higher (P ≤ 0.05) A. marginale parasitemia in animals of T02 than in those of T01. Animals of T01 required one enrofloxacin treatment to decrease A. marginale parasitemia, while those from T02 needed five treatments. From D5 to D37 of study, the mean values of PCV for calves from T02 were lower (P ≤ 0.05) than that for calves from T01. In conclusion, bovines co-infected T. vivax needed four more treatments with enrofloxacin to reduce A. marginale parasitemia and keep PCV values within reference standards.

Spatial distribution of Toxoplasma gondii in cows and associated risk factors.

The aim of the present study was to evaluate the spatial distribution of the prevalence of T. gondii in cows using the indirect immunofluorescence assay and determine associated risk factors. Serum samples were collected from 2970 cows on 263 rural farms in 223 municipalities. A questionnaire was administered to herd owners to collect data for the evaluation of risk factors associated with this disease. Mean seroprevalence of T. gondii in cows was 8.48% (95% CI: 7.48 to 9.49). The microregions with the greatest likelihood (p ≤ 0.05) of having infected animals were Anápolis, Ceres, São Miguel do Araguaia, the Federal District, Anicuns, and Vão do Paraná. The purchase of females or males for reproductive/breeding purposes was significantly associated (p ≤ 0.05) with the prevalence of T. gondii in these regions. A positive correlation (0.7618; p = 0.047) was found between the prevalence of T. gondii and total area in hectares of forests in these regions, suggesting that wild cats may be disseminating T. gondii at these sites. The present results highlight the importance of considering the meat from these animals to be an important infection route for humans who eat raw or undercooked food.

In vitro test for the evaluation of the efficacy of topical products for the control of Cochliomyia hominivorax larvae.

Cochliomyia hominivorax larvae cause myiasis in animals and humans. To register a commercial product to control this dipteran is necessary to experiment on animals. The in vitro test was standardized to evaluate the larvicidal efficacy of commercial topical products. Five formulations were analysed in vitro and in vivo. For the in vitro test, a colony was formed and three replicates (n = 200) of each larval stage (L1, L2 and L3) were treated. The viability of the larvae was evaluated after 5 and 30 min, and at 1, 2, 6, 12, 24, 48, 60 and 72 h post-treatment (HPT). For the in vivo test, 30 bovines divided into six groups were castrated to achieve natural infestation with C. hominivorax. Animals in the treated groups received the product. Myiasis and efficacy were evaluated 12, 24, 36, 48, 60 and 72 HPT. Four formulations tested in the in vitro test achieved 100% efficacy at 24 HPT. In the in vivo experiment only one achieved 100% efficacy at 24 HPT. However, all products achieved the maximum efficacy by the end of study. The in vitro test developed here could be adopted to evaluate the efficacy of topical products for the control of C. hominivorax larvae.

Epidemiological and molecular identification of Trypanosoma vivax diagnosed in cattle during outbreaks in central Brazil.

Bovine trypanosomosis has been spreading in Brazil. In the present study, we evaluated the spatial distribution, prevalence and risk factors of this disease in the state of Goiás, Brazil, and performed both molecular and phylogenetical analyses of Trypanosoma vivax. A total of 4049 blood samples were collected from cattle for a period of 2 years. The parasitological diagnosis was performed using the Woo method and a questionnaire was administered to the farmers to document risk factors associated with the disease in the herd. Positive samples were DNA sequenced and compared to GenBank codes. The prevalence of T. vivax was 8.84%, occurring on 24 ranches only in dairy cattle and mainly in the central and southern portions of the state. The acquisition of new animals infected with T. vivax and the administration of exogenous oxytocin to cows using the same syringe and needle were the main associated factors (P ≤ 0.05). After an outbreak, milk production decreased by 39.62%. The presence of biting flies (tabanids, Haematobia irritans and Stomoxys calcitrans) was not a risk factor (P > 0.05) for the occurrence of T. vivax. The epidemiological data demonstrate the importance of restricting the practice of auctions as well as eliminating the use of exogenous oxytocin in animals during milking. The samples tested by polymerase chain reaction were positive for T. vivax and were genetically homologous with T. vivax found in different states of Brazil and west Africa based on the 18S rRNA gene.

Infection capacity of Trypanosoma vivax experimentally inoculated through different routes in bovines with latent Anaplasma marginale.

The aim of the present study was to compare the infection capacity of Trypanosoma vivax experimentally inoculated through different routes in calves naturally infected with latent Anaplasma marginale. On Day 0 of the study, 25 calves (breed: Girolando) were divided into five groups. The first four groups of five calves each received approximately 1 × 106 trypomastigotes of T. vivax through the intradermal, subcutaneous, intramuscular and intravenous routes. Another five animals remained unaffected to serve as A. marginale naturally infected controls. The study of T. vivax was performed on all calves from D+1 to D+30 using the Woo, Brener and blood smear techniques. PCR was performed on Days +1, +3, +4, +5, +28, +29 and + 30. The results indicated that T. vivax was capable of infecting and developing the disease in the calves independent of the inoculation route. A positive correlation was found between T. vivax and rectal temperature (P ≤ 0.05) and a negative correlation was seen between the protozoan and globular volume (P ≤ 0.05). Latent A. marginale in the calves acted as co-infection for T. vivax. Jaundice was seen only in calves with a high parasitemia by A. marginale. Therefore, in areas with the confirmed presence of T. vivax in bovines, this protozoan should be included in the complex denominated "Bovine Parasitic Sadness", which currently encompasses only Anaplasma and Babesia.

Biological parameters for Rhipicephalus microplus in the field and laboratory and estimation of its annual number of generations in a tropical region.

The present study aimed to evaluate biological parameters of Rhipicephalus microplus in the non-parasitic phase in both field and laboratory conditions. It also aimed to assess correlations between duration (in days) of these parameters and climatic variables (humidity, rainfall, and soil temperature) and to estimate the annual number of generations of R. microplus in a tropical region. The non-parasitic phase of R. microplus in field and laboratory conditions was evaluated throughout the course of two years. A pasture was infested with engorged female of R. microplus, and biological parameters, including female pre-oviposition, female oviposition, egg mass incubation, larval pre-hatching phase, larval maturation, and larval longevity, were evaluated concomitantly with the collection of data on climatic conditions. The same parameters were also evaluated in a climatized chamber in the laboratory. The total duration of the non-parasitic phase in the field was longer in the dry season (1st and 4th life-cycle repetitions) than in the rainy season (2nd, 3rd, and 5th repetitions). Tick biological parameters for the non-parasitic phase in the laboratory were similar to those obtained in the field during the rainy season. The evaluated biological parameters were influenced mainly by environmental and ground-level temperatures, which modified egg mass incubation, larval pre-hatching, and larval longevity periods and, consequently, the total duration of the non-parasitic phase of the tick. The annual number of generations for the tick was estimated at five per year, which is alarming because it represents an increase, and so new studies into strategic control are needed.

Cutting Edge: IgE Plays an Active Role in Tumor Immunosurveillance in Mice.

Exogenous IgE acts as an adjuvant in tumor vaccination in mice, and therefore a direct role of endogenous IgE in tumor immunosurveillance was investigated. By using genetically engineered mice, we found that IgE ablation rendered mice more susceptible to the growth of transplantable tumors. Conversely, a strengthened IgE response provided mice with partial or complete resistance to tumor growth, depending on the tumor type. By genetic crosses, we showed that IgE-mediated tumor protection was mostly lost in mice lacking FcεRI. Tumor protection was also lost after depletion of CD8(+) T cells, highlighting a cross-talk between IgE and T cell-mediated tumor immunosurveillance. Our findings provide the rationale for clinical observations that relate atopy with a lower risk for developing cancer and open new avenues for the design of immunotherapeutics relevant for clinical oncology.

17ß-estradiol differently affects osteogenic differentiation of mesenchymal stem/stromal cells from adipose tissue and bone marrow.

Adipose-derived and bone marrow stem/stromal cells (ASCs and BMSCs) have been often compared for their application in regenerative medicine, and several factors sustaining their differentiation and efficacy have been investigated. 17 ß-estradiol (E2) has been reported to influence some functions of progenitor cells. Here we studied the effects of 10 and 100nM E2 on ASC and BMSC vitality, proliferation and differentiation towards osteogenic and adipogenic lineages. E2 did not modulate ASC and BMSC vitality and growth rate, while the hormone produced a pro-adipogenic effect on both mesenchymal stem/stromal cells (MSCs). In particular, the synergy between 7-day pre-treatment and 100nM E2 led to the most evident result, increasing lipid vacuoles formation in ASCs and BMSCs of +44% and +82%, respectively. Despite the fact that E2 did not alter collagen deposition of osteo-induced MSCs, we observed a different modulation of ASC and BMSC alkaline phosphatase (ALP) activity. Indeed, this osteogenic marker was always enhanced by 17 ß-estradiol in BMSCs, and 7-day pre-treatment with 100nM E2 increased it of about 70%. In contrast, E2 weakened ASC osteogenic potential, reducing their ALP activity of about 20%, with the most evident effect on ASCs isolated from pre-menopausal women (-30%). Finally, we identified an estrogen receptor α (ERα) variant of about 37kDa expressed in both MSCs. Interestingly, adipogenic stimuli drastically reduced its expression, while osteogenic ones mildly increased this isoform in BMSCs only. In conclusion, E2 positively affected the adipogenic process of both MSCs while it favored osteogenic induction in BMSCs only, and both mesenchymal progenitors expressed a novel 37kDa ER-α variant whose expression was modulated during differentiation.

Successful application of virtual screening and molecular dynamics simulations against antimalarial molecular targets.

The main challenge in the control of malaria has been the emergence of drug-resistant parasites. The presence of drug-resistant Plasmodium sp. has raised the need for new antimalarial drugs. Molecular modelling techniques have been used as tools to develop new drugs. In this study, we employed virtual screening of a pyrazol derivative (Tx001) against four malaria targets: plasmepsin-IV, plasmepsin-II, falcipain-II, and PfATP6. The receiver operating characteristic curves and area under the curve (AUC) were established for each molecular target. The AUC values obtained for plasmepsin-IV, plasmepsin-II, and falcipain-II were 0.64, 0.92, and 0.94, respectively. All docking simulations were carried out using AutoDock Vina software. The ligand Tx001 exhibited a better interaction with PfATP6 than with the reference compound (-12.2 versus -6.8 Kcal/mol). The Tx001-PfATP6 complex was submitted to molecular dynamics simulations in vacuum implemented on an NAMD program. The ligand Tx001 docked at the same binding site as thapsigargin, which is a natural inhibitor of PfATP6. Compound TX001 was evaluated in vitro with a P. falciparum strain (W2) and a human cell line (WI-26VA4). Tx001 was discovered to be active against P. falciparum (IC50 = 8.2 µM) and inactive against WI-26VA4 (IC50 > 200 µM). Further ligand optimisation cycles generated new prospects for docking and biological assays.

Human Adipose-Derived Stem Cells on Rapid Prototyped Three-Dimensional Hydroxyapatite/Beta-Tricalcium Phosphate Scaffold.

In the study, we assess a rapid prototyped scaffold composed of 30/70 hydroxyapatite (HA) and beta-tricalcium-phosphate (ß-TCP) loaded with human adipose-derived stem cells (hASCs) to determine cell proliferation, differentiation toward osteogenic lineage, adhesion and penetration on/into the scaffold.In this in vitro study, hASCs isolated from fat tissue discarded after plastic surgery were expanded, characterized, and then loaded onto the scaffold. Cells were tested for: viability assay (Alamar Blue at days 3, 7 and Live/Dead at day 32), differentiation index (alkaline phosphatase activity at day 14), scaffold adhesion (standard error of the mean analysis at days 5 and 18), and penetration (ground sections at day 32).All the hASC populations displayed stemness markers and the ability to differentiate toward adipogenic and osteogenic lineages.Cellular vitality increased between 3 and 7 days, and no inhibitory effect by HA/ß-TCP was observed. Under osteogenic stimuli, scaffold increased alkaline phosphatase activity of +243% compared with undifferentiated samples. Human adipose-derived stem cells adhered on HA/ß-TCP surface through citoplasmatic extensions that occupied the macropores and built networks among them. Human adipose derived stem cells were observed in the core of HA/ß-TCP. The current combination of hASCs and HA/ß-TCP scaffold provided encouraging results. If authors' data will be confirmed in preclinical models, the present engineering approach could represent an interesting tool in treating large bone defects.