Negative Control Research Articles

Green Synthesis of Silver Nanoparticles with Roasted Green Tea: Applications in Alginate–Gelatin Hydrogels for Bone Regeneration

The incorporation of silver nanoparticles (AgNPs) into alginate–gelatin (Alg-Gel) hydrogels can enhance the properties of these materials for bone regeneration applications, due to the antimicrobial properties of AgNPs and non-cytotoxic concentrations, osteoinductive properties, and regulation of stem cell proliferation and differentiation. Here, the hydrogel formulation included 2% (w/v) sodium alginate, 4 µg/mL AgNPs, and 2.5% (w/v) gelatin. AgNPs were synthesized using a 2% (w/v) aqueous extract of roasted green tea with silver nitrate. The aqueous extract of roasted green tea for AgNP synthesis was characterized using HPLC and UHPLC-ESI-QTOF-MS/MS, and antioxidant capacity was measured in Trolox equivalents (TE) from 4 to 20 nmol/well concentrations. Stem cells from human exfoliated deciduous tooth cells were used for differentiation assays including positive (SHEDs/hydrogel with AgNPs) and negative controls (hydrogel without AgNPs). FTIR was used for hydrogel chemical characterization. Statistical analysis (p < 0.05, ANOVA) confirmed significant findings. Roasted green tea extract contained caffeine (most abundant), (−)-Gallocatechin, gallic acid, and various catechins. XRD analysis revealed FCC structure, TEM showed quasispheroidal AgNPs (19.85 ± 3 nm), and UV–Vis indicated a plasmon surface of 418 nm. This integration of nanotechnology and biomaterials shows promise for addressing bone tissue loss in clinical and surgical settings.

Investigating Flavonoids by HPTLC Analysis Using Aluminium Chloride as Derivatization Reagent.

This is the first study to report on high performance thin layer chromatography (HPTLC) generated spectrophotometric data to systematically capture flavonoid compounds using optimized derivatization with either AlCl3 or NaNO2-AlCl3-NaOH as visualisation reagents. While the traditional AlCl3 colorimetric method using UV-Vis analysis provides valuable insights into the presence of flavonoids and allows derivation of the total flavonoid content (TFC) of a sample, HPTLC fingerprints obtained after spraying with AlCl3 or NaNO2-AlCl3-NaOH enable the visualization of the various flavonoids present in a sample based on their respective absorption shifts, thus complementing the traditional TFC assay. In this study, 40 different flavonoids representing different classes (flavonols, flavanolols, flavan-3-ol, flavones, flavanones, and isoflavonoids) were analysed. Upon derivatization with AlCl3 most of the investigated flavonoids recorded bathochromic shifts, yielding characteristic λmax values between 370 and 420 nm, while spraying with NaNO2-AlCl3-NaOH triggered hyperchromic shifts, and thus an increase in absorbance intensity in flavonoids with particular substitution patterns. A few non-flavonoid components with structural similarities to flavonoids (e.g., rosmarinic acid, gallic acid, aspirin, salicylic acid) served as the negative control in this study to determine whether the derivatization reagents allowed exclusive detection of flavonoids. The method was then applied to the analysis of flavonoid containing supplements as well as red clover honey to demonstrate the method's application in the analysis of natural products.

Differences in Anti-inflammatory Effects of Celecoxib Emulgel Preparations with Variations in Surfactant Concentration between Lecithin and Tween 80

Celecoxib is a non-steroidal anti-inflammatory drug (NSAID) which is widely used to treat inflammation such as rheumatoid arthritis and primary dysmenorrhea. The absorption of celecoxib in the body is very low when taken orally, so it can be increased by changing the route of administration via transdermal route such as emulgel. This study aims to determine the increase in the anti-inflammatory activity of celecoxib emulgel with varying concentrations of surfactants in formula with Lecithin and formula with Tween 80. Celecoxib emulgel, was made into 3 formulas with variations in the concentration for each formula with Lecithin (FI=3%; FII=5%; FIII=7%) and Tween 80 (FI=5%; FII=10%; FIII=15%). The anti-inflammatory activity test was carried out by inducing edema in the soles of rats' feet using carrageenan. Rats were divided into 5 groups for each formula: 1) negative control, 2) without surfactant, 3) FI, 4) FII, and 5) FIII. The edema volume was measured every 1 hour for 6 hours using a plethysmometer. Results of the anti-inflammatory test showed that the average value of AUC0-360 in the Lecithin group = 57.97-77.17 μg/hour/mL and in the Tween 80 group = 51.45-77.17 μg/hour/mL. %DAI results of formula without surfactant=9.33%, FI (Lecithin=10.01%; Tween 80=24.49%), FII (Lecithin=18.36%; Tween 80=29.05%), and FIII (Lecithin=24.88%; Tween 80=33.33%). Based on these data, it can be concluded that there is an increase in the anti-inflammatory effect of celecoxib emulgel with increasing surfactant concentration in the formula with Lecithin and in the formula with Tween 80. Keywords: Anti-inflammatory; celecoxib; emulgel; surfactant

Allelopathic interactions of Carthamus oxyacantha, Macrophomina phaseolina and maize: Implications for the use of Carthamus oxyacantha as a natural disease management strategy in maize.

Fungicides are used to control phytopathogens but all these fungicides have deleterious effects. Allelopathic interactions can be harnessed as a natural way to control the pathogens but there are no reports that show the allelopathic interactions of donor plant, recipient crop, as well as the target plant pathogen and the material used for inoculum production. So, in the present study, the suitability of Carthamus oxyacantha M. Bieb. was assessed against Macrophomina phaseolina, the cause of charcoal rot in maize. Among the various treatments in pot experiment, a negative control, 3 concentrations of inoculum (1.2×105, 2.4×105, and 3.6×105 colony forming units (CFU) mL-1, 3 concentrations (0.5, 1.0, and 1.5% w/w) of C. oxyacantha along with an autoclaved M. phaseolina (Mp) and C. oxyacantha alone were included to investigate their allelopathic effects on maize, not investigated earlier. Maximum suppression of the disease was observed by 1.5% (w/w) concentration of C. oxyacantha. Soil amendment with C. oxyacantha significantly suppressed the disease incidence (DI) and disease severity index (DSI) in charcoal rot of maize up to 40 and 55%, respectively over the strongest level of inoculum (Mp3). C. oxyacantha not only reduced area under disease incidence progress curve (AUDIPC) and area under disease severity progress curve (AUDSPC), but also improved the morphological, biochemical and physiological parameters of maize. The maximum increase of 48, 65, and 75% in values of shoot length (SL), shoot dry mass (SDM), and root dry mass (RDM), respectively was observed by application of the highest concentration of C. oxyacantha in the treatment Mp1+Co3, over infested control (Mp1). Photosynthetic pigments, such as chlorophyll a, chlorophyll b and carotenoids were increased to 58, 64, and 46%, respectively over Mp1, by the application of C. oxyacantha. Carbon assimilation rate (A), stomatal conductance (gs), rate of transpiration (E), and internal carbon dioxide concentration (Ci) were significantly increased to 58, 48, 48, and 20%, respectively over infested control (Mp3), by application of C. oxyacantha concentration 1.5 (w/w). Moreover, defense enzymes like superoxide dismutase (SOD), peroxidase (POD) and catalase (CAT) activities were boosted up to 27, 28, and 28% over Mp3, respectively. Positive allelopathy of C. oxyacantha towards maize and negative allelopathy towards M. phaseolina makes C. oxyacantha a suitable candidate for charcoal rot disease control in maize.

Antiinflamation Activity Test of Keji Beling (Strobilanthes crispa (L.) Blume) Leaves Extraction on White Mencites (Mus musculus)

Background: Inflammation often occurs in According to data from WHO, 2004) the number of people with bone inflammation worldwide reached around 11.9 million people. In high-income countries, around 1.3 million people experience bone pain, while in low to middle-income countries, the prevalence reached 5.9 million. In Southeast Asia, there are 4.4 million people with arthritis. In North Sumatra, the prevalence of arthritis according to the diagnosis of health workers is 8.4%, while based on diagnosis or symptoms it reaches 19.2%. In Medan City, the prevalence based on the diagnosis of health workers is 5.1%, and based on diagnosis and symptoms it is 17.2%. The method used in this study is the formation of artificial edema on the soles of the feet of male white mice with 1% carrageenan induction. Testing the anti-inflammatory activity of keji beling leaf extract was carried out with 25 test animals divided into 5 treatment groups. The groups consisted of positive controls given sodium diclofenac at a dose of 6.5 mg/kgBW, negative controls given Na-CMC 0.5%, and three extract groups at doses of 100 mg/kgBW, 200 mg/kgBW, and 300 mg/kgBW. The results showed that the percentage of inflammation in the five test groups decreased consistently from the 60th minute to the 360th minute after carrageenan induction. The highest percentage of inflammation was recorded at the 120th minute for the Na-CMC group, followed by the 100 mg/kgBW, 200 mg/kgBW, 300 mg/kgBW, and sodium diclofenac groups. The highest percentage of inflammation inhibition was obtained from the sodium diclofenac portion, followed by 300 mg/kgBW, 200 mg/kgBW, and 100 mg/kgBW. This shows that the good inhibition percentage is owned by EDKB 300 mg/kgBW continued sodium diclofenac, and EDKB 200 mg/kgBW and 100 mg/kgBW. This indicates that the sodium diclofenac portion and extract with doses of 100, 200, and 300 mg/kgBW have the power to function as anti-inflammatory agents, compared to the Na-CMC group which showed no effect.

Genotoxic and Cytotoxic Activities of Cornhusk Extract of Zea mays and Leaf Extract of Sacharum officinarum

Zea mays husk and Saccharum officinarum have been used for years in ethnomedicine for their antimalarial, anti-inflammatory, antipyretic, antidiabetic, and antiphlogistic activities. The cytotoxic and genotoxic effects of Zea mays husk and Saccharum officinarum leaf extracts on the root meristem cells of Allium cepa were investigated. Onion bulbs were exposed to 2.5 mg/ml, 5mg/ml, and 10 mg/ml concentrations of the extracts for macroscopic and microscopic analysis. Tap water was used as a negative control and Methotrexate (0.1 mg/ml) was used as a positive control. There was statistically significant (p < 0.05) inhibition of root growth depending on concentration by the extracts when compared with the negative control group. All the tested extracts were observed to have cytotoxic effects on cell division in A. cepa. The extract induced chromosomal aberrations and micronuclei (MNC) formations in A. cepa root tip cells were significant (p<0.05) when compared with the control group. The extracts treatment further induced cell death, ghost cells, cells membrane damage, and binucleated cells. The Zea mays husk extract was found to exhibit higher cytotoxic and genotoxic potential than Saccharum officinarum leaf extract. These results suggest that Zea mays husk and Saccharum officinarum leaf extracts possess cytotoxic and genotoxic effects on A. cepa.

Development of a porcine model of skin and soft-tissue infection caused by Staphylococcus aureus, including methicillin-resistant strains suitable for testing topical antimicrobial agents.

In view of the ever-increasing representation of Staphylococcus spp. strains resistant to various antibiotics, the development of invivo models for evaluation of novel antimicrobials is of utmost importance. In this article, we describe the development of a fully immunocompetent porcine model of extensive skin and soft tissue damage suitable for testing topical antimicrobial agents that matches the real clinical situation. The model was developed in three consecutive stages with protocols for each stage amended based on the results of the previous one. In the final model, 10 excisions of the skin and underlying soft tissue were created in each pig under general anesthesia, with additional incisions to the fascia performed at the base of the defects and immediately inoculated with Staphylococcus aureus suspension. One pig was not inoculated and used as the negative control. Subsequently, the bandages were changed on Days 4, 8, 11, and 15. At these time points, a filter paper imprint technique (FPIT) was made from each wound for semi-quantitative microbiological evaluation. Tissue samples from the base of the wound together with the adjacent intact tissue of three randomly selected defects of each pig were taken for microbiological, histopathological, and molecular-biological examination. The infection with the inoculated S. aureus strains was sufficient during the whole experiment as confirmed by both FPIT and from tissue samples. The dynamics of the inflammatory markers and clinical signs of infection are also described. A successfully developed porcine model is suitable for invivo testing of novel short-acting topical antimicrobial agents.

Effect of Melia azedarach seed mediated nano-ZnO on growth performance, protein utilisation efficiency, haematology and nutritional status in pigs

The current study was conducted to investigate the effect of Melia azedarach seed-mediated ZnO nanoparticles on growth performance, protein utilisation efficiency, haematology and nutritional status in pigs. A total of 48 pigs were allocated to the following six treatments replicated 8 times: Negative Control (NC, No antibiotic), Treatment 2: Positive control (PC) given a conventional antibiotic (Oxytetracycline, 40 mg/kg feed); Treatment 3: Nano-ZnO 300 mg/L (N300ZnO), Treatment 4: Group given 150 mg/L Melia azedarach seed mediated nano-ZnO (N150MA), Treatment 5: Group given 300 mg/L Melia azedarach seed mediated nano-ZnO (N300MA), Treatment 6: Group given 450 mg/L Melia azedarach seed mediated nano-ZnO (N450MA). The experiment was conducted over 7 weeks. Melia azedarach seed-mediated ZnO nanoparticles had no significant effect on growth performance apart from average daily feed intake (ADFI) with treatment 3 having the highest value. It significantly affected protein consumption and growth efficiency but not protein efficiency ratio and specific growth rate. Melia azedarach seed-mediated ZnO nanoparticles had no significant impact on nutritional parameters, serum minerals apart from phosphorus which can negatively affect renal functioning.

Formulasi dan Evaluasi Ekstrak Aseton Kulit Putih Semangka Merah Non Biji (Citrullus Vulgaris S) dengan Niacinamide sebagai Gel Pelembab Kulit dengan Uji Kelembapan

This research was carried out with the aim of identifying the content of phytochemical compounds of alkaloids, flavonoids, tannins and saponins in the acetone extract of seedless red watermelon rind (Citrullus vulgaris), knowing the results of evaluating the physical properties of the acetone extract gel of seedless red watermelon rind (Citrullus vulgaris) with niacinamide against various concentrations of moisturizing gel preparations, and comparing the best moisture test concentration on acetone extract gel preparations of seedless red watermelon rind (Citrullus vulgaris) in providing optimal results. This research is a laboratory study of the experimental formulation of acetone extract of seedless red watermelon rind (Citrullus vulgaris) to determine the formulation and concentration of the best extract produced. The research results showed that acetone extract from the white skin of red seedless watermelon (Citrullus vulgaris) contained phytochemical compounds alkaloids, flavonoids, and polyphenol as well as lycopene in the white skin of red seedless watermelon (Citrullus vulgaris). The results of the evaluation of the physical properties of the acetone extract gel of seedless red watermelon rind (Citrullus vulgaris) in the negative control, 2.5% concentration, 5% concentration, and 7.5% concentration met the test requirements but did not meet the spreadability test requirements. And the best concentration of water content test in the acetone extract gel preparation of seedless red watermelon rind (Citrullus vulgaris) in providing optimal results is found in the gel preparation with a concentration of 7.5%.

Antibody titer after anti-idiotype rabies vaccination with nano-chitosan adjuvant

Background.The rabies virus neutralizing antibodies titers is a public health problem in the world, including Indonesia. Rabies is zoonotic and causes death in humans with a case fatality rate of 100%. Anti-idiotype antibody (Ab2) from chicken immunoglobulin (IgY) can be a substitute antigen for the rabies virus. This research aims to study the potential of rabies vaccine based on anti-idiotype antibody (Ab2) with nano-chitosan as an adjuvant.Materials and methods.The production of Ab2 is derived from purified and characterized chicken immunoglobulins. Nano-chitosan is made from chitosan from shrimp shell waste. Vaccination tests were carried out on rats compared with commercial vaccines as a positive control and physiological solutions as a negative control. The characterization results of IgY Ab2 were IgY rabies with BM ~180 kDa, heavy chains (BM ~60 kDa), and light chains (BM ~30 kDa) of IgY. Nano-chitosan is less than 100 nm in size, can dissolve well, and does not cause side effects in experimental animals.Results.The vaccine formula uses a concentration of 0.5%, where the ratio of nano-chitosan and Ab2 is 1:1. The Ab2 concentration used was about 1000 units per mL of the Ab2 suspension.Conclusion.This study concludes that anti-idiotype antibodies dissolved in nano-chitosan adjuvant can induce the formation of antibodies with titers that are not statistically different from the antibody titers induced by commercial rabies vaccines. Nano-chitosan has a potential vaccine adjuvant candidate, safe to use, and can enhance the immunogenicity of an antigen applied subcutaneously. According to the results of this study, it is recommended that post-vaccination antibody titers be measured regularly: for example, every one week for six weeks after vaccination. This measurement determines the time of the peak of the antibody titer so that the time for revaccination can be determined.

A multi-energy inertia-based coordinated voltage and frequency regulation in isolated hybrid power system using PI-TISMC

This paper proposes novel multi-energy inertia support for simultaneous frequency and voltage control of an isolated hybrid power system (IHPS). Multi-energy storage (gas inertia – hydrogen storage, thermal inertia – solar thermal storage, hydro inertia – gravity hydro storage, chemical inertia – battery energy storage) supported by demand side management (DSM) for simultaneous voltage and frequency regulation and backed by biodiesel generators, are the essential elements of IHPS. A novel control strategy of concurrent virtual droop control, virtual damping control, virtual inertia control, and virtual negative inertia control is proposed to utilise multiple inertia sources and to improve LFC and AVR performance effectively. The effective coordination of inertia sources in eradicating oscillations in IHPS, is aided by a developed cascaded proportional integral-tilt-integral-sliding mode (PI-TISMC) controller. The performance of PI-TISMC is compared with PID, PI-PID, and PI-SMC controllers. A maiden attempt has been done by training five diverse classes of optimization techniques to optimize the parameters of controllers in the present work. The results are evaluated in MATLAB and it is evident from the results that the performance of frequency control is improved by 6.5%, 7.8% and 3.4 s (over shoot, undershoot, and settling time). The performance of frequency control is improved by 6.5%, 7.8% and 3.4 s (over shoot, undershoot, and settling time). Similarly, the performance of voltage control is improved by 6.7%, 4.8% and 2.3 s (over shoot, undershoot, and settling time) by employing developed PI-TISMC controller and proposed concurrent inertia control. The combination exhibits superior performance in minimizing oscillations in IHPS due to variations in loading and solar insolation.

Equine bone marrow-derived mesenchymal stromal cells reduce established S. aureus and E. coli biofilm matrix in vitro.

Biofilms reduce antibiotic efficacy and lead to complications and mortality in human and equine patients with orthopedic infections. Equine bone marrow-derived mesenchymal stromal cells (MSC) kill planktonic bacteria and prevent biofilm formation, but their ability to disrupt established orthopedic biofilms is unknown. Our objective was to evaluate the ability of MSC to reduce established S. aureus or E. coli biofilms in vitro. We hypothesized that MSC would reduce biofilm matrix and colony-forming units (CFU) compared to no treatment and that MSC combined with the antibiotic, amikacin sulfate, would reduce these components more than MSC or amikacin alone. MSC were isolated from 5 adult Thoroughbred horses in antibiotic-free medium. 24-hour S. aureus or E. coli biofilms were co-cultured in triplicate for 24 or 48 hours in a transwell plate system: untreated (negative) control, 30 μg/mL amikacin, 1 x 106 passage 3 MSC, and MSC with 30 μg/mL amikacin. Treated biofilms were photographed and biofilm area quantified digitally. Biomass was quantified via crystal violet staining, and CFU quantified following enzymatic digestion. Data were analyzed using mixed model ANOVA with Tukey post-hoc comparisons (p < 0.05). MSC significantly reduced S. aureus biofilms at both timepoints and E. coli biofilm area at 48 hours compared to untreated controls. MSC with amikacin significantly reduced S. aureus biofilms versus amikacin and E. coli biofilms versus MSC at 48 hours. MSC significantly reduced S. aureus biomass at both timepoints and reduced S. aureus CFU at 48 hours versus untreated controls. MSC with amikacin significantly reduced S. aureus biomass versus amikacin at 24 hours and S. aureus and E. coli CFU versus MSC at both timepoints. MSC primarily disrupted the biofilm matrix but performed differently on S. aureus versus E. coli. Evaluation of biofilm-MSC interactions, MSC dose, and treatment time are warranted prior to testing in vivo.

Mushroom and Kefir Functional Characterizations: Hypolipidemia and Gut Microbiota Modulations in Rat Models

ABSTRACTHyperlipidemia is a malnutrition disease associated with different lifestyle factors mainly high fat/cholesterol foods consumption and less physical activity. Consumption of high fiber foods (prebiotic sources) additionally to gut microbiota (GM; probiotics species) could overcome hyperlipidemia and its associated risks. Prebiotics and probiotics are known by protective effects in different diseases like diabetes, inflammatory bowel diseases, and cardiovascular diseases. Mushroom and kefir milk (KM) are known for their high pre/probiotic nutritional values depending on many factors, for example, eating levels and/or conditions. Therefore, this study aimed to measure the potential health benefits of dried powdered mushrooms (DPM) supplemented with KM between hyperlipidemia rats in association with lipid profile, atherogenic index (AI), and GM profile. Rats were randomly divided into main negative control healthy group (G1; −ve), positive control hyperlipidemia (G2; +ve) and three hyperlipidemia groups (G3:G5) fed DPM at 2.5%, 5%, and 10% of rats' diet additionally to 5% KM (DPM + 5% KM) each, respectively. The collected blood samples used for glucose, lipid profile (cholesterol, triglycerides, high‐density lipoprotein cholesterol, and low‐density lipoprotein cholesterol), and AI in addition to fecal sample for GM and short‐chain fatty acids (SCFAs). Collected data illustrated that body weight, blood glucose, lipid profile, GM (Bifidobacteria and Lactobacillus vs. Clostridium histolyticum), AI, and SCFAs were improved between hyperlipidemia fed 5% both PDM + KM (p ≤ 0.05) at the best levels. In conclusion, same DPM/KM levels have broad development as functional active foods that could lower hyperlipidemia incidence and promotes intestinal health; however, much more human studies are needed.

Effects of Whole Flaxseed in Reduced Energy Diets on Broiler Chickens Meat Quality and Carcass Characteristics Under Heat Stress Condition

This study aimed to assess the impact of heat stress (HS) on broiler chickens provided with a low-energy diet including whole flaxseed, focusing on carcass characteristics and meat quality. A total of 250 one-day-old Ross 308 broiler chicks were randomly allocated into five groups (50 chicks per group), with two replicates for each group. The negative control treatment (T1) received a basal diet devoid of HS. The second treatment fed a basal diet under HS as a positive control (T2). In the third experiment, they fed a basal diet, including whole flaxseed, under heat-stress conditions (T3). The fourth treatment consisted of providing a low-energy diet with whole flaxseed under HS (T4). The fifth intervention was a reduced energy diet administered under HS (T5). The carcass characteristics and edible entrails in the T3 and T4 groups showed a significant improvement (P≤0.05) compared to the control groups. The meat quality in the T3 and T4 groups was considerably enhanced (P≤0.05) due to an increase in omega-3 fatty acid levels [α-linolenic acid, eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA)], as determined by GC mass spectrometry compared to the control group that had a corn-soybean diet. In conclusion, incorporating whole flaxseeds into a low-energy diet during HS augmented broiler chickens’ productivity, improved carcass characteristics, and elevated omega-3 concentrations in the meat.

Evaluation of wound healing activities of Vitex doniana (Black Plum) ethanol fruit extract

In this study, the folkloric use of Vitex doniana fruit extract for the treatment of wounds was evaluated. The mesocarps of the fresh fruits were removed, oven-dried at 40°C and pulverized. The powdered material (800 g), was exhaustively macerated in 80 % ethanol, filtered and dried in a water bath at 40°C. Phytochemical screening and acute dermal toxicity of the extract were investigated. Herbal ointments (10 and 50 %w/w) were formulated using soft paraffin base. The wound-healing effect of the extract was carried out using twenty albino rats divided into four groups (n=5). Groups I (soft paraffin) and II (10 %w/w Povidone-iodine) served as negative and positive controls, respectively. Groups III and IV were topically treated with 10 %w/w and 50 %w/w of the herbal ointment respectively. Treatments and wound grading were assessed once daily and every 4 days, respectively, as well as the timeframe for percentage wound contraction and epithelialization Both the 10 %w/w and 50 %w/w herbal ointments exhibited a concentration-related significant (P&lt;0.05) wound contraction when compared to the control. Epithelialization period of 50 %w/w extract was less than that of the standard drug. The wound closure area of 50 %w/w extract was &gt;positive control &gt;10 %w/w extract &gt; negative control. Histological study of the wounds on the 16th day indicated no sign of dermal toxicity. Some secondary metabolites such as alkaloids and tannins were present. In conclusion, the folkloric use of the fruit extract of V. doniana for the treatment of wounds may therefore be justified

Standardization of criteria for interpreting the results of mutagenicity assessment in the Ames test

Introduction. Currently, various combinations of criteria for interpreting the results are used to make a conclusion about the mutagenic activity in the Ames test. In some cases where weak effects are present, especially when evaluating the mutagenicity of generic pesticides that may contain mutagenic impurities, different conclusions are possible, depending on the criteria chosen. Material and methods. To standardize the criteria for interpreting the results in the Ames test, data obtained earlier in the assessment of the mutagenic activity of technical pesticide products were used. The studies were carried out in accordance with the OECD Standard Protocol No. 471 and State Standard (Russian: ГОСТ) 32376–2013 by direct application to the plate and under pre-incubation conditions. Results. The applicability of three combinations of criteria for interpretation of Ames test results was evaluated using our own previously obtained experimental data. It was established that as criteria of biological significance of the results of mutagenicity evaluation in the Ames test it is reasonable to use not only the conservative approach based on the fold increase rule, but also to compare them with the data of the ranges of historical negative laboratory control. Limitations. The study is limited to evaluating the results of experiments obtained using the standard plate test, but not the fluctuation format. Conclusion. A conclusion about the presence of mutagenic activity of the test item in the Ames test can be made if the following criteria are fulfilled: the presence of a statistically significant increase in the number of revertants on the plates with the test item compared to the concomitant negative control; the presence of a concentration-effect relationship; the mean number of revertants for at least one of the concentrations tested, with or without metabolic activation, must exceed the upper limit of the distribution of the historical negative laboratory control; the number of revertants on the plates with the test item must be 2 or more times compared to that one in the negative control for TA97, TA98, TA100, TA102 and 3 or more for TA1535; the reproducible effects.

Study of acute and subacute toxicities and genotoxic and mutagenic potentials of the lyophilized extract of Campomanesia sessiliflora (O.Berg) mattos leaves in wistar rats

Campomanesia sessiliflora (O.Berg) Mattos is a Brazilian native plant species used in a popular medicinal tea for treating gastrointestinal, urinary, and dermatological pathologies. This study evaluated the toxicity of Campomanesia sessiliflora (O.Berg) Mattos via acute and subacute toxicity tests. It also analyzed mutagenic and genotoxic potentials by the micronucleus test, which detects genetic material damage indicating mutagenicity, and the comet assay, which assesses DNA damage levels as a genotoxicity indicator. The plant extract initially originated from the ultrasonic maceration of Campomanesia sessiliflora (O.Berg) Mattos leaves in a hydroethanolic solution. The involved animals were adult Wistar rats. Ten females were available to evaluate acute toxicity and estimate the LD50, receiving a dose of 2000 mg/kg. The subacute toxicity evaluation used 35 females and 35 males divided into seven groups: negative control (saline control – SC), positive control (cyclophosphamide control – CC), 125 mg/kg (125), 250 mg/kg (250), 500 mg/kg (500), 1000 mg/kg (1000), and the satellite group (ST). Genotoxicity and mutagenicity experiments applied bone marrow micronucleus and comet assays. Acute and subacute toxicity tests did not present behavioral, physical, and physiological changes (p≥0.05). Administering the Campomanesia sessiliflora (O.Berg) Mattos extract reduced spleen size in male and female animals, without histopathological changes. However, doses above 500 mg/kg showed significant genotoxic and mutagenic effects in the comet and micronucleus assays compared to the control group. The extract did not exhibit acute or subacute toxicity, but doses higher than 500 mg/kg indicated some level of genotoxicity and mutagenicity.

Hydrogen Gas Inhalation Alleviates Airway Inflammation and Oxidative Stress on Ovalbumin-Induced Asthmatic BALB/c Mouse Model

Airway inflammatory diseases, such as asthma, are a global public health concern owing to their chronic inflammatory effects on the respiratory mucosa. Molecular hydrogen (H2) has recently been recognized for its antioxidant and anti-inflammatory properties. In this study, we examined the therapeutic potential of H2 in airway inflammation using an ovalbumin (OVA)-induced BALB/c mouse model of allergic asthma. Female BALB/c mice were sensitized and challenged with OVA to induce airway inflammation, and 30 mice were randomly divided into five groups: NT (non-treatment), HTC (3% H2 treatment only), NC (negative control, OVA only), PC (positive control, OVA + intranasal 1 mg/mL salbutamol 50 μL), and HT (H2 treatment, OVA + inhaled 3% H2). Various inflammatory and oxidative stress (OS)-induced markers such as white blood cells (WBCs) and their differential counts, lung histology, cytokine levels such as interleukin (IL)-4, (IL)-5, (IL)-13, interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), granulocyte-macrophage colony-stimulating factor (GM-CSF), (IL)-10, reactive oxygen species (ROS), nitric oxide (NO), glutathione peroxidase (GPx), and catalase (CAT), and total immunoglobulin E (IgE) levels were investigated. Our results showed that inhaled H2 significantly reduced inflammatory cell infiltration, OS markers, and pro-inflammatory cytokine expression while upregulating antioxidant enzyme activity. Furthermore, H2 also significantly decreased serum IgE levels, a marker of allergic inflammation. Collectively, our findings suggest that H2 inhalation is a promising treatment option for airway inflammation, offering a novel approach with potential clinical applications.

Antibacterial Activity Of Staphylococcus aureus From Chinese Betel Leaf Juice (Peperomia pellucida L.)

Infectious diseases, especially in developing countries such as Indonesia, are a common health problem, one of which is caused by bacteria. This study tested the antibacterial activity of Chinese betel leaf extract against Staphylococcus aureus bacteria, with secondary metabolite content such as alkaloids, tannins, and flavonoids that act as antibacterials. This study was a laboratory experiment using a completely randomized design with three repetitions at five concentrations of extract, as well as one positive and negative control. The method used was the agar diffusion method, and the test results showed that Chinese betel leaf extract had antibacterial potential against Staphylococcus aureus at various concentrations tested. The concentrations used were 6µL/disc, 8µL/disc, 10µL/disc, 12µL/disc, and 14µL/disc, where all these categories produced inhibition zones included in the moderate category according to Susanto (2012). Statistical analysis using the Kruskal-Wallis test produced a significant value of 0.035 (&lt;0.05), indicating a significant difference between each concentration tested. From the research results, it can be concluded that Chinese betel leaf extract has effective antibacterial activity against Staphylococcus aureus.

MiR-126 accelerates renal injury induced by UUO via inhibition PI3K/ IRS-1/ FAK signaling induced M2 polarization and endocytosis in macrophages

To investigate the role and molecular mechanism of miR-126 in unilateral ureteral occlusion (UUO). We used bioinformatics to analyse miRNAs specifically expressed in UUO. The mouse model of UUO was established using RAW264.7 cells cultured in vitro and in vivo. The mice were divided into control group, miR-126-NC (negative control) group and miR-126-KD (knockdown) group. Then the relative expression of miR-126 was detected by quantitative reverse transcription-polymerase chain reaction (qRT-PCR), the renal fibrosis was detected by Masson staining, and the protein expression of CD68, collagen I and collagen III in the kidney was detected by immunofluorescence assay. Immunohistochemistry detects α-SMA expression. Moreover, Western blotting was performed to measure the expressions of p-PI3K, CD163, CD206, CD86, iNOS, IL-1β, p-FAK, p-Rac-1, p-IRS-1 and MMP9. The relative fluorescence intensity of F-actin and p-FAK was detected by immunofluorescence assay, and the phagocytosis ability of macrophages was determined by phagocytosis assay with fluorescent microspheres. Bioinformatics analysis reveals miR-126-specific overexpression in UUO. Successful transfection of miR-126-NC and miR-126-KD was confirmed by RT-PCR. The selective reduction of miR-126 was validated by Masson, immunohistochemistry and immunofluorescence staining to decrease the area of UUO-induced renal fibrosis and to lower the expression of CD68, α-SMA, collagen I, and collagen III. The reduction of iNOS expression may also be achieved with selective knockdown of miR-126, as verified by cell tests. enhances the phagocytic ability of macrophages and the expression of p-PI3K, CD206, p-FAK, F-actin, p-Rac-1, p-IRS-1 and MMP9. MiR-126 can inhibit the PI3K signaling pathway, promote M1 macrophage polarization, and suppress the activation of FAK and Rac-1, thus accelerating the progression of UUO.