Potential effect of methanol leaf extracts from three Podocarpus species on experimental cryptosporidiosis

@#Cryptosporidiosis causes diarrhea in both immunocompetent and immunocompromised individuals, with acute manifestations occurring particularly in children and the elderly. Up till now, there is no curative therapy for cryptosporidiosis, so discovery of new classes of drugs are of great importance. This study aimed to examine the effect of methanol leaves extracts of the three Podocarpus species; P. macrophyllus (Thunb.), P. gracilior (Pilg.) and P. elongatus (Aiton) L’ Hér. ex Pers and their combination on Cryptosporidium parvum (C. parvum) in experimentally infected mice in comparison with the commercially used drug, Nitazoxanide. As well as spectrophotometric estimation of the total phenolic and flavonoid content of these extracts was done. Results revealed that treatment with these three Podocarpus extracts and their combination showed a significant reduction of the number of C. parvum oocyst shed in the stool of infected mice compared to infected control group and Nitazoxanideinfected treated group at P < 0.001. The combination of the three Podocarpus extracts was the most effective treatment showing the lowest number of oocysts shedding in comparison with other used extracts and Nitazoxanide. Histopathological inspection of sections from ilium and colon displayed signs of improvement after treatment with P. macrophyllus and P. gracilior extracts and more remarkable improvement when the three extracts were combined. It was concluded that the three Podocarpus species extracts used in this study had a promising anti-Cryptosporidium activity especially when they were combined.

Determinación de una nueva constante para la estimación del filtrado glomerular en pediatría / Determination of a new constant to estimate glomerular filtration rate in pediatrics

Introducción. En pediatría, el filtrado glomerular (FG) se puede calcular con el clearance (depuración) de creatinina medida corregida en ml/min/1,73 m2, o se puede estimar según la fórmula de Schwartz (FGe = talla/creatinina plasmática x k). La constante k depende del método de determinación de creatinina plasmática: k = 0,55 para el método colorimétrico de Jaffe, y k =0,413 para el método enzimático. Nuestro laboratorio utiliza el método colorimétrico cinético compensado (MCCC), se observan discordancias entre el FG estimado y el medido.Hipótesis: Los valores de k propuestos no se ajustan al MCCC de creatinina plasmática. Objetivo. Calcular el valor de k que permita estimar el FG mediante la cuantificación de la creatinina con el MCCC. Métodos. Diseño descriptivo correlacional. Se incluyeron pacientes de entre 3 y 18 años con FG normal o alterado atendidos en el Servicio de Nefrología Infantil entre julio de 2017 y enero de 2018 con control de esfínteres y firma del consentimiento. Se excluyeron pacientes desnutridos y con mielomeningocele. Las variables estudiadas fueron: creatinina plasmática y urinaria, talla y diuresis de 24 horas. Resultados. Se analizaron 184 pacientes, con una edad media de 10 años. La mediana del clearance de creatinina medido corregido fue de 123 ml/min/1,73 m2. La correlación lineal entre la talla y la creatinina plasmática y el clearance de creatinina medido corregido arrojó un valor de k de 0,499 (r = 0,974 y r2 = 0,949). La correlación lineal entre el FG estimado (k = 0,499) y el clearance de creatinina medido corregido mostró un coeficiente b = 0,999 (r = 0,951 y r2= 0,903). Conclusión. Según este estudio, la constante que permite estimar el filtrado glomerular al cuantificar la creatinina plasmática con el método colorimétrico cinético compensado es de 0,499.

Introduction. In pediatrics, glomerular filtration rate (GFR) may be estimated by measured corrected creatinine clearance (mcCrCl) (mL/min/1.73 m2) or the Schwartz formula (eGFR = height/plasma creatinine x k). The constant k depends on the plasma creatinine determination method: k = 0.55 for the Jaffe colorimetric method and k = 0.413 for the enzymatic method. Our laboratory uses the compensated kinetic colorimetric assay (CKC), and differences are observed between the estimated and measured GFR.Hypothesis: The proposed values of k do not adjust to the CKC method for plasma creatinine. Objective. To calculate a k value that allows to estimate GFR through creatinine measurement with CKC. Methods. Correlational, descriptive design. Patients aged 3-18 years seen at the Division of Pediatric Nephrology between July 2017 and January 2018 with normal or altered GFR, bladder and bowel control, and signed consent were included. Malnourished and myelomeningocele patients were excluded. Studied variables were plasma and urine creatinine, height, and 24-hour urine output. Results. A total of 184 patients were analyzed, their mean age was 10 years. Median mcCrCl was 123 mL/min/1.73 m2. The linear correlation between height and plasma creatinine and mcCrCl resulted in a k value of 0.499 (r = 0.974 and r2 = 0.949). The linear correlation between the estimated GFR (k = 0.499) and mcCrCl resulted in a 0.999 ß coefficient (r = 0.951 and r2 = 0.903). Conclusion. According to this study, the constant that allows to estimate GFR when measuring plasma creatinine with the CKC method is 0.499.

Partial purification and characterization of antimicrobial peptide(s) from Mimosa pudica (Mimosaceae)

@#Plants have been a major source of natural products for sustaining human health. The use of the different parts of the plant as infusions, decoctions, extracts, and powders are being employed in the treatment of different diseases in humans, plants, and animals. One property of great significance in terms of therapeutic treatments, especially with the emergence of multi-drug resistant microbes, is the antimicrobial activity. A new promising source of antimicrobials that demonstrate novel mechanisms of therapeutic strategies is low molecular weight peptides. In this study, the antimicrobial activities of Mimosa pudica crude and partially purified peptide extracts against Gram-negative Enterobacter cloacae ATCC 23355 and Enterobacter aerogenes ATCC 13048, and Gram-positive Staphylococcus epidermidis ATCC 12228 using resazurin colorimetric assay and tricine SDS-PAGE bioautography were reported. M. pudica crude and partially purified extracts exhibited antimicrobial activity against all the bacteria tested. Specifically, the peptide that was partially purified from M. pudica with a molecular weight of 5.14 kDa inhibited the growth of Enterobacter cloacae.

Inhibition effects of fungal immunomodulatory proteins from Ganoderma spp .on different gastric cancer cell lines in vit ro / 药学实践杂志

Objective To study the cell proliferative effects of fungal immunomodulatory proteins from Ganoderma spp . on 26 gastric cancer cell lines in vitro .Method 26 human gastric cancer cell lines were treated with FIPs by MTS assay .The average optical density (OD) in 490 nm and inhibition rate (GI50 )was counted by Universal Microplate Spectrophotometer . Results Three FIPs showed similar profiling in 26 human gastric cancer cell lines after 72 h treatment in cell proliferation as-say ,which except for NUGC-4 and OCUM-1 did not showed obvious anti-proliferative effect ,the other 24 human gastric cell lines showed some anti-proliferative effects ,especially for 7 cell lines(NUGC-3 ,GTL-16 ,HGC-27 ,IM95m ,SNU-638 ,SNU-216 and SNU-5) showing strong potency ,with their GI50 less than 50 μg/ml .Conclusion FIPs showed strong anti-prolifera-tive effects in some human gastric cancer cell lines in vitro ,which had potential to be further developed as anti-gastric cancer drugs .

Biological characterization of implant surfaces - in vitro study / Caracterização biológica de superfície de titânio - estudo in vitro

Objective: Evaluate the biological performance of titanium alloys grade IV under different surface treatments: sandblasting and double etching (Experimental surface 1; Exp1, NEODENT); surface with wettability increase (Experimental surface 2; Exp2, NEODENT) on response of preliminary differentiation and cell maturation. Material and method: Immortalized osteoblast cells were plated on Exp1 and Exp2 titanium discs. The polystyrene plate surface without disc was used as control group (C). Cell viability was assessed by measuring mitochondrial activity (MTT) at 4 and 24 h (n = 5), cell attachment was performed using trypan blue exclusion within 4 hours (n = 5), serum total protein and alkaline phosphatase normalization was performed at 4, 7 and 14 days (n = 5). Data were analyzed using one-way ANOVA and Tukey test. Result: The values of cell viability were: 4h: C - 0.32±0.01A; Exp1 - 0.34±0.08A; Exp2 - 0.29±0.03A. 24h: C - 0.43±0.02A; Exp1 - ; 0.39±0.01A; Exp2 - 0.37±0.03A. The cell adhesion counting was: C -85±10A; Exp1- 35±5B; Exp2& - 20±2B. The amounts of serum total protein were 4d: C - 40±2B; Exp1 - 120±10A; Exp2 -130±20A. 7d: C 38±2B; Exp1 - 75±4A; Exp2 -70±6A. 14 d: C - 100±3A; Exp1 - 130±5A; Exp2 - 137±9A. The values of alkaline phosphatase normalization were: 4d: C - 2.0±0.1C; Exp1 - 5.1±0.8B; Exp2 - 9.8±2.0A<. 7d: C -1.0±0.01C; Exp1 - 5.3±0.5A; Exp2 - 3.0±0.3B. 14 d: C - 4.1±0.3A; Exp1 - 4.4±0.8A; Exp2 - 2.2±0.2B. ...

Objetivo: Avaliar o desempenho biológico de ligas de titânio grau IV submetidos a diferentes tratamentos de superfície - jateamento e duplo ataque ácido (Superfície experimental 1; Exp1, NEODENT) e superfície com aumento na molhabilidade (Superfície experimental 2; Exp2, NEODENT) em resposta preliminar de diferenciação e maturação celular. Material e método: Foram plaqueados osteoblastos imortalizados sobre discos de titânio de Exp1 e Exp2 e como controle o poço da placa de cultura sem disco (C). Empregou-se ensaios de viabilidade celular (MTT) em 4 e 24 horas (n = 5), adesão celular em 4 horas (n = 5), dosagem de proteínas totais e fosfatase alcalina normalizada em 4, 7 e 14 dias (n = 5). Os dados foram analisados por ANOVA em fator único seguido de teste de Tukey. Resultado: Os valores de viabilidade celular foram: 4h: C - 0,32±0,01A; Exp1 - 0,34±0,08A; Exp2 - 0,29±0,03A. 24h: C - 0,43±0,02A; Exp1 - 0,39±0,01A; Exp2 - 0,37±0,03A. A contagem de adesão celular foi: C - 85±10A; Exp1 - 35±5B; Exp2 - 20±2B. Os valores de proteínas totais foram: 4d: C - 40±2B; Exp1 - 120±10A; Exp2 - 130±20A. 7d: C - 38±2B; Exp1 - 75±4A; Exp2 - 70±6A. 14 d: C - 100±3A; Exp1 - 130±5A; Exp2 - 137±9A. Os valores de fosfatase alcalina normalizada foram: 4d: C - 2,0±0,1C; Exp1 - 5,1±0,8B; Exp2 - 9,8±2,0A, 7d: C - 1,0±0,01C; Exp1 - 5,3±0,5A; Exp2 - 3,0±0,3B, 14 d: C - 4,1±0,3A; Exp1 - 4,4±0,8A; Exp2 - 2,2±0,2B. Letras diferentes representam ...

Desarrollo y validación de un método de determinación por espectrofotometría visible de antimoniato de meglumina en inyectables / Development and validation of a colorimetric assay for the determination of meglumine antimoniate in injections

En este artículo se presentan los resultados del desarrollo y de la validación de un método analítico para realizar la determinación de antimoniato de meglumina en un inyectable para uso humano. El método se funda en la medida de la absorción a 352 nm de la coloración producida al hacer reaccionar el antimoniato de meglumina con yoduro de potasio en medio ácido. El método validado se desarrolló para hacer la determinación del antimoniato de meglumina como parte del control de calidad y para llevar a cabo estudios de estabilidad de la molécula en el inyectable. Como variables de validación se estudiaron la selectividad, la linealidad, la precisión, la exactitud y la robustez.

In this paper we present the results obtained during the development and validation of an analytical methodology by a colorimetric method for the assay of meglumine antimoniate in injections. The quantification implies the formation of a colored complex resulting after of meglumine antimoniate and potassium iodure in acid media. The absorbance was made at 352 nm. The validated methodology may be used as a part of quality control with release purpose and also, in chemical stability studies of the injections. As validation parameters, we studied selectivity, lineality, precision, accuracy and robustness.

Evaluation of a Colorimetric Broth Microdilution Method for Antimicrobial Susceptibility Testing Using 2,3,5-Triphenyltetrazolium Chloride / 대한임상미생물학회지

BACKGROUND: The broth microdilution susceptibility testing method is considered a standard for determining minimum inhibitory concentrations, and the addition of the redox indicator 2,3,5-triphenyltetrazolium chloride (TTC) to the broth microdilution method simplifies and increases its objectivity. The current study evaluated the usefulness of a TTC-modified broth microdilution method for antimicrobial susceptibility test of frequently encountered clinical isolates. METHODS: The minimal inhibitory concentrations (MICs) of 10 antimicrobials for 111 clinical isolates of four bacterial species, Staphylococcus aureus, Escherichia coli, Enterobacter cloacae, and Acinetobacter baumannii, were investigated by a modification of the Clinical and Laboratory Standards Institute (CLSI)-recommended broth microdilution method with the addition of 2,3,5-triphenyltetrazolium chloride (TTC). The inhibitory effects of TTC against 192 strains of 22 bacterial species isolated from clinical specimens were also evaluated. RESULTS: The number of colonies of all 192 strains of 22 bacterial species grown on TTC-containing Mueller-Hinton agar did not differ from those grown on Mueller-Hinton agar only. The MICs with TTC were within 2 dilutions of those obtained by the CLSI method in 569 (97.6%) of 583 organism-antimicrobial agent combinations. CONCLUSIONS: The colorimetric MIC method using TTC may be a useful surrogate of antimicrobial susceptibility testing for most of the frequently isolated bacteria.

A RAPID COLORIMETRIC ASSAY FOR SURVIVAL AND PROLIFERATION OF BACTERIUM AND FUNGI / 解剖学报

Objective To introduce a rapid colormietric assay for survival and proliferation of bacterium and fungi. Methods Colorimetric assay and automatic microplate scanning spectrophotometer were used for assay of survival and proliferation of bacteria and fungi in the present work. Results Close correlation has been found between the A (570?nm) values of the formazan products and the cell concentration of living bacteria and fungi detected.Conclusion The present work has developed an effective, sensitive and convenient assay method for survival and proliferation assay of bacteria and fungi. [

Inhibitory Effect of Mitomycin C (MMC) on the Proliferation of Human Corneal Keratocyte

PURPOSE: To evaluate the anti-proliferative effect of mitomycin C (MMC) on human corneal keratocyte, and to investigate the cellular morphology of keratocyte according to the concentration and exposure time in vitro. METHODS: Human corneal keratocytes using endothelium-free explant method were exposed to 0.005%, 0.01%, and 0.05% concentration of MMC for 3, 5, and 10 minutes. MTT based colorimetric assay was performed to assess the inhibition of cellular proliferation, and cellular morphology was evaluated by inverted phase-contrast light microscope and electron microscope. RESULTS: Use of higher concentration MMC and prolongation of exposure time resulted in greater inhibitory effect on cellular proliferation. When exposed to 0.005% MMC for 3, 5 and 10 minutes, the survival rate of keratocyte was 100%, 95.7% and 74.0% respectively. At 0.01% MMC, the survival rate was 98.6%, 92.9%, and 66.9%. At 0.05% MMC, it was 74.0%, 73.4%, and 38.8%. Exposure to the highest concentration (0.05%) among the 3 preparations for 3 or 5 minutes showed significant inhibition of keratocyte proliferation (p<0.05), and when exposed for 10 minutes, all 3 preparations showed significant inhibition of keratocyte proliferation (p<0.05). Inverted phase-contrast light microscopy showed that human corneal keratocytes lost their adherence to the bottom of the dish and assumed round and swollen shape rather than spindle shape when exposed to higher concentration of MMC for a prolonged time. The damaged keratocytes showed the degenerative changes like cellular membrane disruption, disappearance of microvilli, enlargement of rough surfaced endoplasmic reticulum and mitochondria, and vacuole formation by electronic microscope. CONCLUSIONS: When MMC is applied to inhibit the proliferation of keratocytes involved in corneal wound healing, it seems to be a valuable application at least 0.05% concentration for 3 minutes. Further studies should be followed for the biological effect of MMC including drug toxicity associated with human corneal tissue in vivo.

Effect of Anti-inflammatory Mediator on the Proliferation of Human Corneal Keratocyte

PURPOSE: The purpose of this study was to evaluate the effect of anti-inflammatory mediators like dexamethasone, nordihyroguaratic acid (NDGA), and diclofenac sodium on proliferation of human corneal keratocytes, and to investigate the cellular morphology of keratocyte. METHODS: Human corneal keratocytes were exposed to 0.05, 0.1, 0.3, 0.8, and 1.0 mM concentration of each drug for period of 24, 48, and 72 hours. MTT based colorimetric assay was performed to assess the metabolic activity and inhibition of cellular proliferation. Cellular morphology was evaluated by inverted phase contrast micrograph and electron microscopy. RESULTS: The higher the concentration of inoculated each drugs was, the more the inhibitory effect of human keratocyte proliferation was found (P<0.05). NDGA, over 0.3 mM and diclofenac, more than 0.1 mM had significant more inhibitory effect on keratocyte proliferation compared with dexamethasone within 48 hours of exposure to each drug. With the concentration and exposure time of each drug, human corneal keratocytes were visible more rounded and swollen rather than spindle shape, and detached from the bottom of the dish. The damaged keratocytes had degenerative changes like cellular membrane disruption, microvilli disappearance, enlarged rough surfaced endoplasmic reticulum and mitochondria, vacuole formation and nuclear membrane damage by TEM. CONCLUSIONS: On basis of this study, the anti-inflammatory mediators such as NDGA and diclofenac sodium have less side effects and stronger inhibitory effects of human keratocyte proliferation than dexamethasone.

Chemosensitivity testing for human gastric cancer with short-term culture cells vs purified primary culture cells using the MTT assay / 中国癌症杂志

Purpose:To evaluate the feasibility, advantages and disadvantages of chemosensitivity test of human gastric cancer using MTT assay with short term culture tumor cells contaminated by nonmalignant cells compared with purifiedly primary culture tumor cells.Methods:Fifty nine fresh samples from patients with gastric cancer were obtained from operating rooms. Chemosensitivity results were provided by the MTT assay with short term culture cells. The primary culture cells were purified by means of a series of methods such as repeated cell attachment, differential trypsinization and natural purification which removed fibroblasts and other nonmalignant cells. The same MTT assay was conducted using purified cells. Chemosensitivity results between short term method and purification method were compared for seven antitumor drugs. Results:The success rate of short term method using the MTT assay for chemosensitivity testing was 81 4% (48 of 59 patients),and for the purification method it was 50.8(30 of 59 patients). The average was 20.2?9.5 days when primary cells were cultured into purified cancer cells. The optical density ( A ) value correlated directly with the the number of tumor cells with the two methods( P

The Effect of Mitomycine C (MMC) on Inhibition of Pterygial Fibroblast Proliferation

The purpose of this study is to investigate that the biological effect of mitomycin C(MMC) in cellular metabolic activity and morphological change on the ptreygium fibroblast in vitro by MMC concentration and duration of exposure used clinically. Human pterygial fibroblasts were exposed for threeminute and five-minute to MMC 0.002%, 0.004%, 0.01%, 0.02%, 0.04%, and DMEM(control). MTT based colorimetric assay was performed to assess the metabilic activity, inhibition of fibroblast proliferation on the MMC concentration and exposure time. The higher the concentration of MMC, and longer the duration of exposure time, the absorbance of spectrometer are decreased. The metabolic activity of fibroblasts were inhibited by 50% at least only over MMC 0.02% for five-minute expoure time. In histological findings, the higher the concentration and longer the duration of MMC exposure time, the enlargement of many mitochondira and rough endoplasmic reticulum without nuclear damage were more distinctly appeared. Especially, the ptergial fibroblast has more severe cytoplasmic damage at a five-minute exposure to MMC 0.02% than a three-minute exposure to MMC 0.04%. For inhibition of fibroblast proliferation, in case of using MMC should be at least over 0.02% concentration for five-minute exposure time. Arthors think that the experimental and clinical research on the duration of MMC exposure time as well as the concentration MMC, should be need to evaluate the effect on inhibition of cellular proliferation.

Application of MTT Colorimetric Assay in Research of Cytokine Activity and Cytotoxicity of Killer Cells / 中国肿瘤生物治疗杂志

We used MTT assay to test the cellular cytotoxicity ( NK, LAK, CTL, Macrophage), cytokine activities ( 1L-1, 1L-2, 1L-6, TNF), proliferation of lymphocytes and chemosensitivity of tumor cells, and compared it with radioactive isotope assay. The results showed that the MTT assay may be used to test the cellular cytotoxicity, cytokine ac-tivity, proliferation of lymphocytes and chemosensitivity of tumor cells. We think it is a simple, rapid, economic and safety method.

In Vitro Effect of Secnidazole Benzoate on Trichomonas vaginalis / 中国寄生虫学与寄生虫病杂志

Objective To research the trichomonacidal effect of secnidazole benzoate in vitro.Methods Trichomonas vaginalis was cultured in liver extract medium in 96-well microplate.The culture suspension of Trichomonas vaginalis was divided into four groups:secnidazole benzoate, secnidazole, metronidazole and control, with medium as blank control.MTT colorimetric assay was applied to determine the inhibitory effect of secnidazole benzoate on the proliferation of Trichomonas vaginalis.The culture suspension was transfered into test tubes and divided into same groups to observe inhibitory effect by the classical microscopic counting method.Results After 24 h incubation, the proliferation of the parasites was concentration-dependent by secnidazole benzoate(t=9.02, P

The regulation mechanism of shRNAspten to NR2B after neuron subjected to anoxia insult / 重庆医科大学学报

Objective:To investigate the regulation of PTEN with RNAi on NR2B after OGD.Methods:In the OGD model with anaerobic gas mixture and deoxygenated,glucose-free extracellular solution,shRNAspten-GFP and unrelated control plasmid PshGFP transfection were done respectively.The quantities of NR2B in the neuronal membrane were quantitatively measured with colorimetric assay,and the expression of NR2B mRNA were analyzed by RT-PCR.The neuronal activity was analyzed by AlamarBlue andintracellular free calcium concentration were measured with Fura-2 AM by laser confocal microscope(LCM).Results:shRNAspten-GFP plasmid can been expressed successfully in cultured neurons.The study demonstrated that there are numerous NR2B subunit expression on neuronal membrane by colorimetric assay(OPD)assay and RT-PCR.The expression and quantity of neuronal NR2B with shRNAspten-GFP treatment was significantly decrease after oxygen and glucose deprivation(OGD)and reperfusion(P