Low prevalence of ideal levels in cardiovascular behavior metrics among Mexican adolescents.

BACKGROUND: Lifestyle changes when transitioning from high-school to college expose students to unhealthy behaviors associated with high cardiovascular risk. The study aimed to assess the cardiovascular behavior metrics according to the AHA criteria, in freshman college adolescents from Northwest Mexico. METHODS: The study was cross-sectional. Demographics and health history were collected by questionnaires. Four behaviors were evaluated: diet quality using a duplicated FFQ, physical activity (PA) using the IPAQ, smoking, and body mass index (BMI) percentile; blood pressure was measured as a biological metric. Intakes were averaged and summed for each food group; sodium and saturated fat were calculated using the Mexican System of Food Equivalents or the USDA Database. Metrics were categorized into ideal, intermediate or poor level according to the AHA criteria. Diet outliers (± 3 SD) were trimmed and data was tested for normality. Mean±SD were calculated for continuous and percentages for categorical variables. Chi-square test compared the prevalence of demographic variables and levels of each cardiovascular metric by sex. Independent T-test evaluated differences in anthropometrics, dietary, and PA by sex, and the prevalence of ideal vs. non-ideal dietary intakes. RESULTS: Participants were n = 228, 55.6% men, age = 18.5±0.4 y. A higher prevalence of men indicated working, playing sports, and family history hypertriglyceridemia (p < 0.05). Men showed higher weight, height, BMI, waist, blood pressure, and lower PA and body fat (p < 0.05). Concerning diet quality, significant differences by sex were observed in nuts and seeds (1.1±0.6 and 0.9±0.6 oz/week, p = 0.042) and processed meats (749.8±639 and 503.6±300.3 g/week, p = 0.002); only fish and shellfish group reached AHA recommendations (513.1 ± 450.7 vs. 501.7 ± 428 g/week, p = 0.671) for men and women, respectively. Ideal level was reached by 70.9% participants for BMI percentile, 87% for smoking, 67.2% for blood pressure, 25.9% for PA, and 12.2% for diet score. Regarding food groups and nutrients, the lower prevalence in the ideal level was for sugar-sweetened beverages (10%, p = 0.013) and processed meats (4.8%, p = 0.208), and the highest for fish and shellfish (87.8%, p = 0.281) . CONCLUSIONS: The diet and PA patterns of Northwest Mexican freshman adolescents make them a high-risk group for developing long-term unhealthy habits and cardiovascular complications early in adulthood.

Nanopharmaceuticals (Au-NPs) after use: Experiences with a complex higher tier test design simulating environmental fate and effect.

The current environmental hazard assessment is based on the testing of the pristine substance. However, it cannot be excluded that (nano)pharmaceuticals are excreted into sewage during the use phase followed by entry into wastewater treatment plants (WWTPs). Sorption to sewage sludge or release via effluent can result in modified ecotoxicological effects which possibly can only be detected with a modified test approach. The objective of our study was to investigate a realistic exposure scenario for metallic nanoparticles (NPs) in pharmaceutical products, excreted into effluent, and released into the environment after treatment in WWTPs. The test approach was illustrated by using gold (Au) NPs. Effluent from model WWTPs were investigated in aquatic tests (Daphnia magna, fish cell lines). Sewage sludge was used as a sole food source (Eisenia fetida) or mixed with soil and used as test medium (soil microorganisms, Folsomia candida, Enchytraeus crypticus). To cover the aspect of regulation, the test systems described in OECD-test guidelines (OECD TG 201, 211, 220, 232, 249, 317) were applied. Modifications and additional test approaches were included to meet the needs arising out of the testing of nanomaterials and of the exposure scenarios. The results were assessed regarding the suitability of the test design and the toxicity of Au-NPs. Except for activated sludge as a sole food source for E.fetida, the selected test approach is suitable for the testing of nanomaterials. Additional information can be gained when compared to the common testing of the pristine nanomaterials in the standardized test systems. Effects of Au-NPs were observed in concentrations exceeding the modeled environmental.

Predictive Factors for Unsuccessful Use of Arteriovenous Fistula in a Population of End-Stage Renal Disease Patients in Southeastern Mexico.

BACKGROUND: An arteriovenous fistula (AVF) is the preferred vascular access for long-term hemodialysis. The main disadvantage of AVF is the rate of nonmaturation or unsuccessful use for hemodialysis (FUHD). We described our findings in AVF creation and possible risk factors associated with FUHD. METHODS: This is a retrospective study of AVFs during a 6-year period. Variables collected at the time of creation were demographics, comorbidities, replacement therapy, preoperative laboratory tests, and estimated 6-month mortality on hemodialysis. All AVFs were created in the upper arms. Outcomes were FUHD, cannulation failure, and cumulative survival. Univariate and multivariate analyses were performed to find possible risk factors for FUHD. RESULTS: AVFs were created in 78 patients. Average age was 36.3 years, and 74.4% were male. Mean body mass index was 24.5 kg/m2. The most common etiologies were glomerulopathy (53.6%) and diabetes mellitus (13.4%). Estimated six-month mortality was 4.2%. One patient underwent AVF before hemodialysis (mean dialysis time 2.2 years). Nineteen AVFs were considered FUHD (23.2%). Cannulation failure was 15.9%. AVF 1-year and 3-year survival was 67.8% and 63.5%, respectively. FUHD had higher estimated six-month mortality on hemodialysis, shorter prothrombin time, and lower serum albumin level than successful AVF (univariate analysis) (P < 0.05) Short prothrombin time and albumin were confirmed for FUHD (multivariate analysis). A 3.3-gr/dL serum albumin cutoff point (area under the curve, 0.715; receiver operating characteristic) (P < 0.05) was determined for FUHD. CONCLUSIONS: The population referred for AVF creation possesses different characteristics in our center. Good AVF outcomes can be achieved. Preoperative serum albumin level and prothrombin time could be the possible risk factors associated with unsuccessful AVF use.

Simulation Studies to Explore Biodegradation in Water-Sediment Systems: From OECD 308 to OECD 309.

Studies according to OECD 308 and OECD 309 are performed to simulate the biodegradation of chemicals in water-sediment systems in support of persistence assessment and exposure modeling. However, several shortcomings of OECD 308 have been identified that hamper data evaluation and interpretation, and its relation to OECD 309 is still unclear. The present study systematically compares OECD 308 and OECD 309 and two variants thereof to derive recommendations on how to experimentally address any shortcomings and improve data for persistence and risk assessment. To this end, four (14)C-labeled compounds with different biodegradation and sorption behavior were tested across standard OECD 308 and 309 test systems and two modified versions thereof. The well-degradable compounds showed slow equilibration and the least mineralization in OECD 308, whereas the modified systems provided the highest degree of mineralization. Different lines of evidence suggest that this was due to increased oxygenation of the sediment in the modified systems. Particularly for rapidly degrading compounds, non-extractable residue formation was in line with degradation and did not follow the sediment-water ratio. For the two more slowly degrading compounds, sorption in OECD 309 (standard and modified) increased with time beyond levels proposed by equilibrium partitioning, which could be attributed to the grinding of the sediment through the stirring of the sediment suspension. Overall, the large differences in degradation observed across the four test systems suggest that refined specifications in test guidelines are required to reduce variability in test outcomes. At the same time, the amount of sediment and its degree of oxygenation emerged as drivers across all test systems. This suggests that a unified description of the systems was possible and would pave the way toward a more consistent consideration of degradation in the water-sediment systems across different exposure situations and regulatory frameworks.

Proteomic profile of the venom of three dark-colored Tityus (Scorpiones: Buthidae) from the tropical rainforests of Costa Rica.

OBJECTIVE: We aimed to elucidate the potential differences in the venom peptide sequences of three Tityus species from Costa Rican rainforests: T. jaimei, T. championi and T. dedoslargos, compared to T. cf. asthenes from Colombia, which could explain the low level of scorpionism in Costa Rica, evidenced by the lack of epidemiological data. METHODOLOGY: We applied venom proteomics of peptides purified by RP-HPLC and compared the obtained sequences from venoms of these Tityus species to the sequences previously identified from Tityus inhabiting other Central and South American regions. RESULTS: Venom proteome analysis evidences that most of the putative peptide toxins identified in Costa Rican dark-colored Tityus are very similar to those present in other T. (Atreus) from the region. CONCLUSIONS: Our study suggests that, in the case of potential envenomation by Tityus in Costa Rica, the same level of toxicity should be observed, compared to other cases caused by members of the subgenus from other geographical localities. On the other hand, compared to countries with more accelerated urban expansion, Costa Rican Tityus still inhabit secondary rainforests and do not commonly share the same spaces with humans, so the lack of epidemiological evidence of medical emergencies caused by envenoming by this scorpion group could be more related to ecological and demographic factors and less attributed to the characteristics of the venom.

Cannabis Sativa targets mediobasal hypothalamic neurons to stimulate appetite.

The neurobiological mechanisms that regulate the appetite-stimulatory properties of cannabis sativa are unresolved. This work examined the hypothesis that cannabinoid-1 receptor (CB1R) expressing neurons in the mediobasal hypothalamus (MBH) regulate increased appetite following cannabis vapor inhalation. Here we utilized a paradigm where vaporized cannabis plant matter was administered passively to rodents. Initial studies in rats characterized meal patterns and operant responding for palatable food following exposure to air or vapor cannabis. Studies conducted in mice used a combination of in vivo optical imaging, electrophysiology and chemogenetic manipulations to determine the importance of MBH neurons for cannabis-induced feeding behavior. Our data indicate that cannabis vapor increased meal frequency and food seeking behavior without altering locomotor activity. Importantly, we observed augmented MBH activity within distinct neuronal populations when mice anticipated or consumed food. Mechanistic experiments demonstrated that pharmacological activation of CB1R attenuated inhibitory synaptic tone onto hunger promoting Agouti Related Peptide (AgRP) neurons within the MBH. Lastly, chemogenetic inhibition of AgRP neurons attenuated the appetite promoting effects of cannabis vapor. Based on these results, we conclude that MBH neurons contribute to the appetite stimulatory properties of inhaled cannabis.

Dual substrate sensor platform for rapid detection and differentiation of coliforms.

In the present work, we modified and improved the previously developed sensor platform (based on screen-printed carbon electrode) for rapid detection and quantification of coliforms. The second substrate (L-alanyl-p-nitroanilide) was included in the sensor platform, which produced the second cathodic voltammetric peak at the new potential (-113 mV) in the defined potential range (-600 to 400 mV). The 106 CFU/mL of 5 different groups of coliforms in 3 h of incubation produced electrochemical signals equal to 960 µA, 970 µA, 988 µA, 950 µA, 956 µA and 920 µA respectively. After 3 h of incubation the two strains of Gram-positive bacteria with concentration of 106 CFU/mL produced voltammetric signals equal to --80 µA and- 200 µA (due to high /-150 µA/ capacitive/background current) respectively. No voltammetric peak occurred with Gram-positive bacteria at the potential equal to -113 mV. The voltammetric signals produced at the new potential range were specific for coliforms.

Intrageneric cross-reactivity of monospecific rabbit antisera against venoms of the medically most important Bitis spp. and Echis spp. African snakes.

BACKGROUND: Snakebite envenomation exerts a heavy toll in sub-Saharan Africa. The design and production of effective polyspecific antivenoms for this region demand a better understanding of the immunological characteristics of the different venoms from the most medically important snakes, to select the most appropriate venom combinations for generating antivenoms of wide neutralizing scope. Bitis spp. and Echis spp. represent the most important viperid snake genera in Africa. METHODOLOGY/PRINCIPAL FINDINGS: Eight rabbit-derived monospecific antisera were raised against the venoms of four species of Bitis spp. and four species of Echis spp. The effects of immunization in the rabbits were assessed, as well as the development of antibody titers, as judged by immunochemical assays and neutralization of lethal, hemorrhagic, and in vitro coagulant effects. At the end of immunizations, local and pulmonary hemorrhage, together with slight increments in the plasma activity of creatine kinase (CK), were observed owing to the action of hemorrhagic and myotoxic venom components. Immunologic analyses revealed a considerable extent of cross-reactivity of monospecific antisera against heterologous venoms within each genus, although some antisera provided a more extensive cross-reactivity than others. The venoms that generated antisera with the broadest coverage were those of Bitis gabonica and B. rhinoceros within Bitis spp. and Echis leucogaster within Echis spp. CONCLUSIONS/SIGNIFICANCE: The methodology followed in this study provides a rational basis for the selection of the best combination of venoms for generating antivenoms of high cross-reactivity against viperid venoms in sub-Saharan Africa. Results suggest that the venoms of B. gabonica, B. rhinoceros, and E. leucogaster generate antisera with the broadest cross-reactivity within their genera. These experimental results in rabbits need to be translated to large animals used in antivenom production to assess whether these predictions are reproduced in horses or sheep.

Comparison of adjuvant emulsions for their safety and ability to enhance the antibody response in horses immunized with African snake venoms.

Adjuvant emulsions are widely used to enhance the antibody response in animals used as immunoglobulin source to produce snake antivenoms. We tested the performance of four commercial emulsion adjuvants (Montanide, Freund, Carbigen, and Emulsigen-D) and an experimental adjuvant (QH-769) in the antibody response of horses towards venoms of the African snakes Bitis arietans, Echis ocellatus, Dendroaspis polylepis and Naja nigricollis. Montanide, Freund and Carbigen adjuvants generated the highest immune response but induced moderate/severe local lesions at the site of injection. In contrast, Emulsigen-D and QH-769 adjuvants generated the lowest immune response and low incidence of local lesions. No evidence of systemic alterations was observed in the horses immunized with any of the adjuvants. It is suggested that the use of Montanide or Freund-based emulsions in the first immunization steps, followed by the use of Emulsigen-D, QH-769 or similar adjuvants in the following injections, could result in a satisfactory immune response against snake venoms, while not inducing serious local deleterious effects.

Importance of Mevalonate Pathway Lipids on the Growth and Survival of Primary and Metastatic Gastric Carcinoma Cells.

PURPOSE: This preclinical study aims to determine the effect of drugs that alter isoprenoids and cholesterol metabolism in the homeostasis of gastric carcinoma cell lines in the search for new therapeutic targets for stomach cancer. MATERIALS AND METHODS: Primary (AGS) and metastatic (NCI-N87) gastric cancer cell lines were treated with simvastatin and terbinafine, two inhibitors of the mevalonate pathway, and avasimibe, an inhibitor of cholesterol esterification. Cell viability and growth were measured as well as cholesterol levels and the expression of the hydroxy methyl-glutaryl CoA reductase (HMGCR) and the LDL receptor (LDLR). RESULTS: Primary and metastatic gastric carcinoma cells show different sensitivity to drugs that affect isoprenoid synthesis and the metabolism and uptake of cholesterol. Isoprenoids are involved in the growth and viability of both types of cells, but the role of free and esterified cholesterol for metastatic gastric cell survival is not as evident as for primary gastric cancer cells. Differential expression of LDLR due to mevalonate pathway inhibition suggests variations in the regulation of cholesterol uptake between primary and metastatic cancer cells. CONCLUSION: These results indicate that at least for primary gastric cancer, statins and avasimibe are promising candidates as potential novel antitumor drugs that target the metabolism of isoprenoids and cholesterol of gastric tumors.

Microbial Population Dynamics in Model Sewage Treatment Plants and the Fate and Effect of Gold Nanoparticles.

Adequate functioning of a sewage treatment plant (STP) is essential to protect the downstream aquatic environment (ECHA 2017), and information on the degradability of chemicals and their toxicity to activated sludge microorganisms is required. An environmental realistic higher tier test is a STP simulation test as described in OECD 303A (2001) which for nanoparticles can also be used to study their sorption behavior to activated sludge. However, information is limited on the influence of synthetic sewage on the microbial community of the activated sludge. A modified community can result in modifications of the sludge floccules affecting the sorption behavior. The main objective of our study was to show whether a representative microbial diversity remains under standardized test conditions as described in OECD 303A (2001) using synthetic sewage as influent. Furthermore, we investigated whether just considering the functional properties of a STP (elimination of dissolved organic carbon; nitrification), is sufficient for an assessment of gold nanoparticles (AuNPs) or whether the influence on microbial diversity also needs to be considered. AuNPs were used as a case study due to their rising medical applications and therefore increasing probability to reach the sewer and STP. The results can provide significant input for the interpretation of results from the regulatory point of view. To deliver these objectives, the general changes of the microbial population in activated sludge and its influence on the degradation activity (dissolved organic carbon (DOC) and inorganic nitrogen) using freshly collected sludge from the municipal STP in an artificial test system as a model STP in accordance with OECD 303A (2001) were assessed. Additionally, we evaluated the potential impact of AuNPs and its dispersant on the microbial composition and the overall impact on the function of the STP in terms of DOC degradation and nitrogen removal to observe if an assessment based on functional properties is sufficient. The bacteria composition in our study, evaluated at a class level, revealed commonly described environmental bacteria. Proteobacteria (ß, α, δ) accounted for more than 50% but also nitrifying bacteria as Nitrospira were present. Our results show that mainly within the first 7 days of an acclimatization phase by addition of synthetic sewage, the bacterial community changed. Even though AuNPs can have antibacterial properties, no adverse effects on the function and structure of the microorganisms in the STP could be detected at concentrations of increased modeled PEC values by a factor of about 10,000. Complementary to other metallic nanomaterials, gold nanomaterials also sorb to a large extent to the activated sludge. If activated sludge is used as fertilizer on agricultural land, gold nanoparticles can be introduced into soils. In this case, the effect on soil (micro)organisms must be investigated more closely, also taking into account the structural diversity.

Expanding the neutralization scope of the Central American antivenom (PoliVal-ICP) to include the venom of Crotalus durissus pifanorum.

The proteomic, enzymatic, toxicological, and immunogenic profiles of the venom of C. d. pifanorum were studied. It was found that venom of C. d. pifanorum is composed of 63% phospholipases A2 (PLA2s), 13% serine proteinases (SVSPs), 8% bradykinin-potentiating peptides (BPPs), 4% L-amino acid oxidases (LAAOs), 3% metalloproteinases (SVMPs), and other minor components. This composition allows the venom to exert lethal, PLA2, myotoxic, coagulant and defibrinogenating activities, but no azocaseinolytic or hemorrhagic activities. The addition of C. d. pifanorum venom to the group of venoms used as immunogens to produce the Central American antivenom PoliVal-ICP (i.e., venoms of Bothrops asper, Crotalus simus and Lachesis stenophrys) resulted in 1) the expansion of the neutralization scope of the antivenom to cover the venom of C. d. pifanorum and other antigenically related venom (i.e., C. s. scutulatus venom), 2) improvement of the neutralizing potency towards the venom of C. simus, and 3) no significant reduction of the neutralization of venoms of B. asper and L. stenophrys. It was concluded that supplementation of the immunogens used to produce PoliVal-ICP with the venom of C. d. pifanorum is a viable alternative to expand the neutralization scope of the antivenom. BIOLOGICAL SIGNIFICANCE: The venom of Crotalus durissus pifanorum from Venezuela has a proteomic profile like those of other subspecies of the South American rattlesnake C. durissus, with predominance of phospholipases A2 (especially crotoxin) and serine proteinases. This explains a toxicological profile characterized by neurotoxicity, myotoxicity, and coagulopathies, but being devoid of hemorrhagic activity. The antivenom used in Central America (PoliVal-ICP) includes the venom of C. simus, which has a different composition, in the immunizing strategy. Accordingly, this antivenom does not neutralize C. d. pifanorum venom. The addition of C. d. pifanorum venom to the immunizing mixture of PoliVal-ICP expands the neutralizing scope of this antivenom, to cover additional rattlesnake venoms, while not affecting the response to C. simus, Bothrops asper and Lachesis stenophrys venoms. This represents an improvement of the current PoliVal-ICP.

Assessing a 6-h endpoint observation time in the lethality neutralization assay used to evaluate the preclinical efficacy of snake antivenoms.

The lethality neutralization assay in mice is the gold standard for the evaluation of the preclinical efficacy and specification fulfillment of snake antivenoms. However, owing to the animal suffering involved, this assay is a candidate to be replaced by in vitro alternatives or, at least, improved by the reduction of the number of animals used per experiment, the introduction of analgesia, and the refinement of the test. Since these tests are usually run for 24 or 48 h, one possibility to refine it is to shorten the endpoint observation time of the assay and so limiting the duration of suffering. To assess the effect of this modification of the standard procedure on the analytical properties of the assay, we compared the median lethal dose (LD50) and median effective dose (ED50) values, estimated through observation times of 6, 24 and 48 h. We used African and Latin American snake venoms and several batches of two polyspecific antivenoms. A significant correlation was found between LD50 and ED50 values estimated at the three observation times. Although some LD50 and ED50 values were significantly different at these time points, results of 6 h were robust enough to be used in the characterization of new antivenoms, the verification of specification compliance, and the parallel comparison of formulations. Our observations support the modification of the standard procedures used for assessing neutralizing ability of antivenoms by carrying out the observations at 6 h instead of 24 or 48 h, with the consequent reduction in the suffering inflicted upon mice during these assays. However, the shortening of the observation time in the lethality tests must be validated for each venom and antivenom before its introduction in the routine procedures.

Prevalence of osteoporosis and osteopenia in a cohort of HIV positive women with a history of treated neoplasms.

INTRODUCTION: Higher prevalence of osteopenia and osteoporosis in HIV positive patients compared to non-infected population has been recognized. However, cancer patients have a higher risk of bone loss and fractures that is multifactorial. The aim of the study was to describe the prevalence of osteopenia and osteoporosis in HIV positive women with history of treated cancer. METHODS: Between January 2018 and December 2019, women aged >40 years, HIV+ with a history of cancer diagnosis, who attended the AIDS Cancer Clinic at Instituto Nacional de Cancerología, Mexico City, and who had a dual X-ray absorptiometry performed during the study period were included. Two control groups (CG)-HIV negative women with history of cancer (CG1) and non-HIV, non-cancer women (CG2)-were matched by age 1:1. RESULTS: Forty-eight patients in each group were included; the mean age was 51.1 ± 8.1 years. Osteopenia was found in femoral neck in 54.2% (HIV+), 37.5% (CG1), and 27.1% (CG2), p = 0.02; in spine was 35.7%, 47.9%, and 31.2%, respectively, p = 0.442. Osteoporosis in femoral neck was documented in 12.5%, 2.1%, and 0% in HIV+, CG1, and CG2 (p = 0.03), and in the spine was 47.9%, 16.7%, and 14.6%, respectively (p = 0.002). CONCLUSION: HIV patients with a history of treated cancer have a much higher prevalence of osteoporosis when compared with same-aged HIV-uninfected women with and without cancer. It is necessary to monitor Bone Mineral Density periodically, and all patients should be encouraged to make lifestyle changes, such as avoid tobacco and alcohol, and to increase exercising.

In vitro Characterization of Anti-SARS-CoV-2 Intravenous Immunoglobulins (IVIg) Produced From Plasma of Donors Immunized With the BNT162b2 Vaccine and Its Comparison With a Similar Formulation Produced From Plasma of COVID-19 Convalescent Donors.

Despite vaccines are the main strategy to control the ongoing global COVID-19 pandemic, their effectiveness could not be enough for individuals with immunosuppression. In these cases, as well as in patients with moderate/severe COVID-19, passive immunization with anti-SARS-CoV-2 immunoglobulins could be a therapeutic alternative. We used caprylic acid precipitation to prepare a pilot-scale batch of anti-SARS-CoV-2 intravenous immunoglobulins (IVIg) from plasma of donors immunized with the BNT162b2 (Pfizer-BioNTech) anti-COVID-19 vaccine (VP-IVIg) and compared their in vitro efficacy and safety with those of a similar formulation produced from plasma of COVID-19 convalescent donors (CP-IVIg). Both formulations showed immunological, physicochemical, biochemical, and microbiological characteristics that meet the specifications of IVIg formulations. Moreover, the concentration of anti-RBD and ACE2-RBD neutralizing antibodies was higher in VP-IVIg than in CP-IVIg. In concordance, plaque reduction neutralization tests showed inhibitory concentrations of 0.03-0.09 g/L in VP-IVIg and of 0.06-0.13 in CP-IVIg. Thus, VP-IVIg has in vitro efficacy and safety profiles that justify their evaluation as therapeutic alternative for clinical cases of COVID-19. Precipitation with caprylic acid could be a simple, feasible, and affordable alternative to produce formulations of anti-SARS-CoV-2 IVIg to be used therapeutically or prophylactically to confront the COVID-19 pandemic in middle and low-income countries.

High Efficacy of Therapeutic Equine Hyperimmune Antibodies Against SARS-CoV-2 Variants of Concern.

SARS-CoV-2 variants of concern show reduced neutralization by vaccine-induced and therapeutic monoclonal antibodies; therefore, treatment alternatives are needed. We tested therapeutic equine polyclonal antibodies (pAbs) that are being assessed in clinical trials in Costa Rica against five globally circulating variants of concern: alpha, beta, epsilon, gamma and delta, using plaque reduction neutralization assays. We show that equine pAbs efficiently neutralize the variants of concern, with inhibitory concentrations in the range of 0.146-1.078 µg/mL, which correspond to extremely low concentrations when compared to pAbs doses used in clinical trials. Equine pAbs are an effective, broad coverage, low-cost and a scalable COVID-19 treatment.

Development and characterization of two equine formulations towards SARS-CoV-2 proteins for the potential treatment of COVID-19.

In the current global emergency due to SARS-CoV-2 outbreak, passive immunotherapy emerges as a promising treatment for COVID-19. Among animal-derived products, equine formulations are still the cornerstone therapy for treating envenomations due to animal bites and stings. Therefore, drawing upon decades of experience in manufacturing snake antivenom, we developed and preclinically evaluated two anti-SARS-CoV-2 polyclonal equine formulations as potential alternative therapy for COVID-19. We immunized two groups of horses with either S1 (anti-S1) or a mixture of S1, N, and SEM mosaic (anti-Mix) viral recombinant proteins. Horses reached a maximum anti-viral antibody level at 7 weeks following priming, and showed no major adverse acute or chronic clinical alterations. Two whole-IgG formulations were prepared via hyperimmune plasma precipitation with caprylic acid and then formulated for parenteral use. Both preparations had similar physicochemical and microbiological quality and showed ELISA immunoreactivity towards S1 protein and the receptor binding domain (RBD). The anti-Mix formulation also presented immunoreactivity against N protein. Due to high anti-S1 and anti-RBD antibody content, final products exhibited high in vitro neutralizing capacity of SARS-CoV-2 infection, 80 times higher than a pool of human convalescent plasma. Pre-clinical quality profiles were similar among both products, but clinical efficacy and safety must be tested in clinical trials. The technological strategy we describe here can be adapted by other producers, particularly in low- and middle-income countries.

Cell death induced by Bothrops asper snake venom metalloproteinase on endothelial and other cell lines.

Two adherent cell lines, BAEC and HeLa, and non-adherent Jurkat, were treated with snake venom metalloproteinase BaP1 to determine whether cytotoxicity, previously reported for this toxin, could be mediated by the process of anoikis. It was observed that there was no correlation between the ability of this toxin to induce loss of adherence, and the cytotoxic effect, since concentrations that do not induce loss of adherence (3-6 microg/mL), were able to trigger 50% of cytotoxicity in BAEC. In the case of HeLa, where toxicity was very low (less than 20% at maximun concentrations and times of exposure), significant detachment and no toxicity was observed at concentrations of 1.5 microg/mL, showing also no correlation between both events. We also observed differences between BAEC toxicity measured by XTT reduction and DNA fragmentation determined by flow cytometry (as an indicator of apoptosis), since concentrations that induce 100% of cytotoxicity barely showed any DNA fragmentation (12% at 24h), suggesting that if apoptosis was involved, DNA damage is still not present, although chromatin condensation, another indicator of apoptosis, is observed in 40% of the cells. Inhibition of BAEC cytotoxicity by caspase inhibitors indicate that apoptosis is playing a role in this process, but other mechanisms of cell death could be participating also. Another way to determine whether the mechanism of cell death was related to anoikis was using a non-adherent cell line, which should show substrate independence. We determined by TUNEL that at 50 microg/ml BaP1 triggered 50% of apoptosis at 96 h, an effect that was seen earlier, suggesting also that if this toxin was inducing apoptosis in a non-adherent cell line, the mechanism could not be related to loss of attachment. Cell cycle arrest in S phase was also observed in Jurkat cells, an effect that could be leading to apoptosis. In conclusion, since there was no correlation between cell detachment and cytotoxicity (and apoptosis) in adherent cell lines and due to the ability of BaP1 to induce apoptosis in a non-adherent cell line, we suggest that this enzyme is toxic by a mechanism not related to anoikis, and that in the case of Jurkat cells, it is likely to be related to its ability to induce cell cycle arrest. Processes other than apoptosis could be also involved in the cell death mechanism mediated by BaP1 on BAEC.

Carotenoid composition and antioxidant activity of the raw and boiled fruit mesocarp of six varieties of Bactris gasipaes.

Total carotenoid content and composition of carotenoids of six varieties of Bactris gasipaes were determined by spectrophotometry and HPLC, with photodiode array detector. Significant differences in total carotenoid content (1.1 to 22.3 mg/100g) were detected among these varieties. Boiling the fruits for 30 minutes did not affect total carotenoid content, but did change the amount of some specific carotenoids, mainly by the production of Z-isomers. Peach palm varieties had the same carotenoids, but in different proportions, presenting mainly, all E-beta-carotene (26.2% to 47.9%), Z-gamma-carotene (18.2% to 34.3%) and Z-lycopene (10.2% to 26.8%). When antioxidant activity was evaluated using DPPH, it was observed that the variety with higher percentages of beta-carotene (54.1%) presented the higher activity. This is one of the first reports in carotenoid content and antioxidant activity in well typified varieties of Bactris gasipaes, whose results could have a positive impact in the consumption of certain peach palm varieties.

Mevalonate pathway as a novel target for the treatment of metastatic gastric cancer.

Gastric mucosa tumors may present as two distinct major entities: Diffuse and intestinal subtypes. There is no standard treatment for advanced or metastatic gastric cancer. The mevalonate pathway and cholesterol homeostasis are important processes in cancer cells that may be highly relevant in terms of cell growth, survival and metastatic potential. Two model cell lines representing intestinal (NCI-N87) and diffuse (Hs746T) metastatic gastric tumor histological subtypes were treated with different drugs that alter membrane lipid metabolism to determine whether cell proliferation, viability and migration were affected. The results indicated that the cells exhibited significant differences in proliferation when treated with the cholesterol-lowering drug simvastatin, but not with terbinafine, another compound that affects cholesterol synthesis. Only simvastatin affected migration in both cell lines. Reposition studies with mevalonolactone, farnesyl pyrophosphate and geranylgeranyl pyrophosphate in the presence of high and low FBS concentrations indicated that both isoprenoids and cholesterol reversed the antiproliferative effects of simvastatin in gastric cancer cells. The cell lines used in the present study had different sensitivities to several potential anti-neoplastic agents that affect the synthesis of membrane lipids. The diffuse gastric cancer cells were particularly sensitive to simvastatin, suggesting it as an option for combination treatment.