Measurement Of Dehydrogenase Activity Research Articles

Modified-release gliclazide acutely improves recovery but causes undesirable blood glucose decrease after a resistance exercise session in healthy adults: a pilot study for a randomized clinical trial.

Sulfonylureas have been used to improve performance in strength sports. However, this hypothetical effect has not been proven. We examined the ergogenic acute effect of gliclazide on resistance training performance and muscle recovery. We conducted a double-blind, randomized, crossover pilot study with 10 healthy resistance-trained adults (29.3 ± 4.4 years), nonusers of anabolic steroids. The participants were randomized to two exercise sessions. In the first session, five participants received placebo and the other five received gliclazide modified release, both administered 8 hours before the session. Session two was performed in a crossover fashion a week later. The volume load was calculated as the maximum number of repetitions of four sets multiplied by load (65% 1-RM). Blood samples were collected before and after exercise, as well as 24 hours and 48 hours after exercise for measurement of creatine kinase (CK-MM) and lactate dehydrogenase (LDH) activity. Blood glucose was measured with a glucometer before, during, and after the exercise sessions. Gliclazide did not enhance volume load for bench press (placebo: 2,698.0 ± 923.0 kg; gliclazide: 2,675.0 ± 1,088.0 kg; p = 0.073) or leg press (placebo: 10,866.0 ± 2,671.0 kg; gliclazide: 10,817.0 ± 2,888.0 kg; p = 0.135). However, CK-MM (-27.7%; p = 0.034) and LDH (-21.1%; p = 0.021) activities were decreased with gliclazide 48 hours after exercise. There was also a decrease in blood glucose in the gliclazide compared with the placebo session (p = 0.018). Gliclazide did not enhance performance in a single resistance training session, but promoted faster muscle recovery. The decrease in blood glucose post-exercise with gliclazide was an undesirable effect that could lead to long-term glucose metabolism disorders. Registered in ClinicalTrials.gov under number NCT04443777.

ALDH Activity Assay: A Method for Cancer Stem Cell (CSC) Identification and Isolation.

Cancer stem cells (CSCs) are a small tumor cell subpopulation, driving cancer initiation, progression, multidrug resistance, and metastasis. Several methods are used to detect and isolate CSCs by flow cytometry. Among these, measurement of aldehyde dehydrogenase (ALDH) activity within the cell is an assay widely used to identify and isolate CSCs from different types of solid tumors. The aldehyde dehydrogenase (ALDH) is a polymorphic enzyme responsible for the oxidation of aldehydes to carboxylic acids, overexpressed both in normal and cancer stem cells. In this chapter, it is described how CSCs are detected and isolated by using ALDH activity assay.

Protective Effect of Dihydromyricetin Against ExerciseInduced Muscle Damage and Its Mechanism

Objective To investigate the protective effect of dihydromyricetin (DHM) against exercise-induced muscle damage (EIMD) in mice and its potential mechanism.Methods Adult male C57BL/6J mice were randomly divided into control group (CG), exercise group (EG), and exercise + 100 mg/kg weight ·d DHM (DHM) group. The intervention lasted for four weeks, during which the animals in the EG and DHM groups were subjected to exercise training for 1 h per day. The day after the training, a 90-min treadmill exercise (slope: 0 and speed: 18 m/min) was conducted in both EG and DHM groups. Samples of blood and gastrocnemius muscles were harvested from the three groups 24 h after the exercise, followed by the measurement of serum creatine kinase (CK) and lactate dehydrogenase (LDH) activities, total superoxide dismutase (T-SOD) activity, malondialdehyde (MDA), and skeletal muscle mitochondrial enzyme complex I and II activities. Histological changes in the skeletal muscle were observed by transmission electron microscopy, and the protein expressions of mitochondrial function-related pathways were detected by Western blotting.Results Skeletal muscle morphological changes and mitochondrial damage were alleviated in the DHM group compared to those in the EG. The activities of EIMD markers CK and LDH and the level of lipid peroxidation were notably repressed and the serum T-SOD activity was enhanced after DHM intervention. Western blotting demonstrated that the expressions of sirtuin type 3 (SIRT3), estrogen-related receptor alpha, and peroxisome proliferator-activated receptor-gamma coactivator-1 alpha in the skeletal muscle of mice increased after the DHM intervention.Conclusion DHM can relieve EIMD in mice, possibly by promoting the recovery of the mitochondrial structure and function in the skeletal muscle of mice after high-intensity exercise via the activation of the SIRT3 signaling pathway.

Hyperpolarized [1-13 C] pyruvate MRSI to detect metabolic changes in liver in a methionine and choline-deficient diet rat model of fatty liver disease.

Nonalcoholic fatty liver disease is an important cause of chronic liver disease. There are limited methods for monitoring metabolic changes during progression to steatohepatitis. Hyperpolarized 13 C MRSI (HP 13 C MRSI) was used to measure metabolic changes in a rodent model of fatty liver disease. Fifteen Wistar rats were placed on a methionine- and choline-deficient (MCD) diet for 1-18 weeks. HP 13 C MRSI, T2 -weighted imaging, and fat-fraction measurements were obtained at 3 T. Serum aspartate aminotransaminase, alanine aminotransaminase, and triglycerides were measured. Animals were sacrificed for histology and measurement of tissue lactate dehydrogenase (LDH) activity. Animals lost significant weight (13.6% ± 2.34%), an expected characteristic of the MCD diet. Steatosis, inflammation, and mild fibrosis were observed. Liver fat fraction was 31.7% ± 4.5% after 4 weeks and 22.2% ± 4.3% after 9 weeks. Lactate-to-pyruvate and alanine-to-pyruvate ratios decreased significantly over the study course; were negatively correlated with aspartate aminotransaminase and alanine aminotransaminase (r = -[0.39-0.61]); and were positively correlated with triglycerides (r = 0.59-0.60). Despite observed decreases in hyperpolarized lactate signal, LDH activity increased by a factor of 3 in MCD diet-fed animals. Observed decreases in lactate and alanine hyperpolarized signals on the MCD diet stand in contrast to other studies of liver injury, where lactate and alanine increased. Observed hyperpolarized metabolite changes were not explained by alterations in LDH activity, suggesting that changes may reflect co-factor depletion known to occur as a result of oxidative stress in the MCD diet. HP 13 C MRSI can noninvasively measure metabolic changes in the MCD model of chronic liver disease.

Evaluating phytochemical constituents and in-vitro antiplasmodial and antioxidant activities of Fadogiella stigmatoloba, Hygrophylla auriculata, Hylodesmum repandum, and Porphyrostemma chevalieri extracts

IntroductionFadogiella stigmatoloba, Hygrophylla auriculata, Hylodesmum repandum and Porphyrostemma chevalieri are used against malaria in traditional medicine in the Democratic Republic of the Congo (DRC). To evaluate their potential in the treatment of this disease, the in vitro antiplasmodial property of these four plants was evaluated. All experiments were conducted on methanolic extracts performed on selected organ parts of these plants. MethodsThe methanolic extracts, obtained by maceration, were firstly screened in vitro against the chloroquine sensitive (3D7) and resistant (W2) Plasmodium falciparum strains by the measurement of lactate dehydrogenase activity, and on human keratinocytes (HaCat) cells by the MTT assay to determine their selectivity indices (SI). Secondly, the antioxidant activity of the same extracts was evaluated using DPPH and FRAP assays. Finally, the presence of specific phytochemical constituents was evaluated using standard methods and tentatively identified by GC-MS. ResultsAn optimum antiplasmodial activity (IC50 = 3.4 ± 0.7 μg/mL, for 3D7, SI = 58.2; IC50 = 7 ± 1.0 μg/mL, for W2, SI = 28.3) was obtained with the leave extract of P. chevalieri. The leaves (for F. stigmatoloba and H. repandum), and the aerial part (for H. repandum) extracts showed promising and moderate antiplasmodial activities against respectively the 3D7 strain (IC50: <15 μg/mL), and W2 strain (IC50:15–50 μg/mL). All extracts presented a weak cytotoxic effect (IC50: >100 μg/mL) on HaCat cells. For the antioxidant test, the most interesting activity was obtained with the leaf extract of P. chevalieri. The GC–MS analysis of these four plants species extracts revealed the presence of various compounds, such as Ethyl 2-nonenoate, 2-(2-Hydroxy-2-phenylethyl)-3,5,6-trimethyl pyrazine, Palmitic Acid, Ethyl palmitate, Ethyl linolenate, and N-Acetyltyramine. ConclusionBased on the obtained results, P. chevalieri could be selected for further investigations or ⁄and for the management of malaria after standardization.

Mammosphere formation assay optimization in the characterization of cancer stem cells of the primary breast tumor

Background/Purpose: Breast cancer is the most frequently diagnosed cancer and the leading cause of death from cancer among females worldwide. Breast cancer tumors that feature breast cancer stem cells (BCSCs) are known to cause drug resistance and metastasis. Culturing BCSCs from primary tumors as mammospheres is both difficult and costly. Therefore, the ability to form BCSC mammospheres in-vitro has become essential in assessing their characterization. Flow cytometric analysis of surface markers and measurement of aldehyde dehydrogenase (ALDH) activity are among other methods used to evaluate cancer cells' stem cell activity. &#x0D; Methods:The research material consisted of BCSCs isolated from the tumor tissues collected from two patients with invasive ductal carcinoma breast tumors. Subsequently, several stem cell surface markers, i.e., cluster of differentiation 44 (CD44), CD24, and CD133, were analyzed using flow cytometry during the third passaging of the cells. ALDH assay is performed with negative control verapamil incubated cells. Two mammosphere forming methods, i.e., low attachment and agar-coated wells together with medium seeded in three different cell concentrations, were compared. &#x0D; Results: CD44+, CD24- and CD133+antibody expressions showed that these cells could be tumor-initiating CSCs. ALDH assay results also indicated that these cells possessed stem cell features. In addition, the results of the mammosphere assay revealed that agar-coated wells at a concentration of 7000 cells/cm2 had more prominent floating features and viable spheres.&#x0D; Conclusion: The findings of this study supported the hypothesis that agar-coated culture dishes in mammosphere culture would increase the mammosphere formation efficiency (MFE) value and revealed the importance of the number of cells in elucidating the nature of BCSCs.

NISIN and gilaburu (Viburnum opulus L.) combination is a cost-effective way to control foodborne Staphylococcus aureus

This study aims to elucidate the inhibitory activity of nisin alone or in combination with gilaburu ( Viburnum opulus L.) against the foodborne Staphylococcus aureus . The effect of nisin and gilaburu on the growth of S. aureus was investigated under different pH (5–7) and salt concentrations (NaCl, 0.5–4.5% w/v) at 30 °C. Nisin exhibited the best antibacterial activity at pH 5 with a minimum inhibitory concentration (MIC) of 25 μg/mL, while its MIC was 50 μg/mL in all salt concentrations when the pH was kept constant at pH 7. The MIC of gilaburu was found to be 50% in all conditions studied. The minimum bactericidal concentrations (MBC) of nisin and gilaburu were 100 μg/mL and >50%, respectively. Changes in pH and salt concentrations did not affect MBC. According to checkboard assay results, a synergy was found between gilaburu and nisin (FIC = 0.25) for both 2.5% and 4.5% w/v salt concentrations at pH 5. The synergistic interaction between nisin and gilaburu was also studied using an agar disk diffusion assay where nisin concentration was kept constant at 3.125 μg/ml, while gilaburu concentration varied between 3.125 and 100%. The positive effect of gilaburu on nisin activity was further confirmed by time-kill assay, flow cytometry, and measurement of relative TTC dehydrogenase activity and conductivity as well as DNA and protein leakage. In conclusion, nisin combined with gilaburu exerted the same inhibitory effect on S. aureus at a concentration approximately 8 times lower than that of MIC of nisin alone (25 μg/mL). This study shows the potential use of nisin and gilaburu in combination to control S. aureus in food. • S. aureus can be controlled with 8 times less nisin in the presence of gilaburu. • Synergy was found between gilaburu and nisin (FIC = 0.25) according to checkboard assay. • Synergistic effect of nisin and gilaburu on S. aureus was proven by different methods. • Nisin and gilaburu combination is an effective strategy to control S. aureus in food.

Comparison of Analytical Values after Changing to the International Standardized Method for Lactate Dehydrogenase and Alkaline Phosphatase Measurements in Mouse and Rat.

Since April 2020, the method for lactate dehydrogenase (LD) and alkaline phosphatase (ALP) activity measurements in Japan has been switched from the Japan Society of Clinical Chemistry (JSCC) reference method, which is only used in Japan, to the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) reference method. However, in some species, the relationship between the blood values of both enzymes measured by the two methods remains unclear. Hence, values measured by these two methods cannot be used interchangeably. Therefore, this relationship was examined in ICR mice and Wistar/ST rats. The LD and ALP values obtained by both methods were plotted on scatter graphs, and regression equations were obtained. To compare the JSCC (x) and IFCC (y) methods, regression equations were generated for LD values in non-hemolytic samples as follows: y = 0.954x - 4.008 for ICR mice and y = 0.963x - 6.324 for Wistar/ST rats. The conversion factors from the JSCC to the IFCC methods were 0.954 (mice) and 0.963 (rats). The conversion coefficients from the IFCC to the JSCC methods were 1.048 (mice) and 1.088 (rats). For ALP values in fasted mouse and rat samples, the regression equations were y = 0.336x - 2.247 and y = 0.314x - 17.626, respectively. The conversion factors from the JSCC to the IFCC methods were 0.336 (mice) and 0.314 (rats). The conversion coefficients from the IFCC to the JSCC methods were 2.978 (mice) and 3.188 (rats). These conversion factors can be used for the mutual conversion of both measured values during the transition period from the JSCC to the IFCC method. However, it should be noted that the conversion coefficients for both LD and ALP were affected by isozyme composition.

Protective Effects of One 2,4-Dihydro-3H-Pyrazol-3-one Derivative against Posterior Capsular Opacification by Regulation of TGF-β2/SMADs and Non-SMAD Signaling, Collagen I, and Fibronectin Proteins.

Many elderly individuals frequently experience cataracts that interfere with vision. After cataract surgery, the left lens epithelial cell (LEC) exhibited fibrosis and posterior capsule opacification (PCO). Sometimes, there is a need for a second surgery; nevertheless, people try other methods, such as a good pharmacological agent, to treat PCO to reduce transforming growth factor-β2 (TGF-β2) amounts to avoid secondary surgery. The aim of the present study was to explore the potential anti-PCO activity of five 2,4-dihydro-3H-pyrazol-3-one (DHPO) derivatives in a TGF-β2-induced fibrogenesis SRA01/04 cell model. The 2-phenyl-5-propyl-DHPO (TSE; no. 2: TSE-2) compound showed the best activity of reduced expression levels of TGF-β2 among five derivatives and therefore was chosen to evaluate the anti-PCO activity and molecular mechanisms on the Sma and mad protein (SMAD) signaling pathway (including TGF-β2, SMADs, and the inhibition of nuclear translocation of SMADs), non-SMAD pathway proteins, including p-extracellular, regulated protein kinases (ERK) 1/2, or p-c-Jun N-terminal kinase (JUN) by Western blotting, PCR, or confocal immunofluorescence analyses. Following treatment with 10 μg/mL of the five compounds, the cells displayed great viability by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTT) assay. In this study, the result of lactate dehydrogenase (LDH) activity measurement did not affect the cytotoxicity of the five compounds. In TGF-β2-induced fibrogenesis in SRA01/04 cells, treatment with the TSE compound decreased the TGF-β2/SMAD signaling genes, including reduced mRNA or expression levels of TGF-β2, SMAD3, and SMAD4, leading to inhibition of TGF-β2-induced fibrogenesis. Our confocal immunofluorescence analyses demonstrated that TSE treatment displays a suppressive effect on SMAD2/3 or SMAD4 translocation to the nucleus. Furthermore, TSE treatment exhibits a reduction in the non-SMAD target gene expression levels of p- c-Jun N-terminal kinase (JUN), p- extracellular, regulated protein kinases (ERK)1/2, p- p38 mitogen-activated protein kinase (p38), p-phosphatidylinositol 3-kinase (PI3K), p-mammalian target of rapamycin complex (mTORC), p-Akt (Ser473), and p-Akt (Thr308). The overall effect of TSE is to reduce the expression levels of collagen I and fibrinogen (FN), thus contributing to antifibrotic effects in cell models mimicking PCO. Our findings reveal the benefits of TSE by regulating TGF-β/SMAD signaling and non-SMAD signaling-related gene proteins to display antifibrotic activity in cells for the possibility of preventing PCO after cataract surgery.

ФЕНОМЕН ПРОТИВОЛУЧЕВОЙ ЗАЩИТЫ ИНДРАЛИНОМ КРУПНЫХ ЖИВОТНЫХ (СОБАК) И ЕГО ЭКСТАПОЛЯЦИЯ НА ЧЕЛОВЕКА

Testing the effectiveness of radioprotectors in large animals is an important stage in the introduction of drugs into the practice of medicine. Unique in scale and world practice, comprehensive studies of the radioprotector of emergency action of the drug B-190 (indralin) in experiments on dogs were carried out. Its high radioprotective efficiency was established with gamma, gamma-neutron and proton high energy irradiation. Under gamma irradiation of dogs indralin at an optimal dose of 30 mg/kg had DRF equel 3, the result of which was not recorded in large animals by any of known radioprotectors. It is important that under non-uniform radiation (shielding of abdomen, head or pelvis), protective effect of indralin doubles. The drug is also active in peroral use. Analysis of extrapolation of experimental data on radioprotective properties of indralin from large animals (dogs) to humans on measurement of succinate dehydrogenase (SDH) activity in lymphocytes of peripheral blood is carried out. Activation of SDH during stress response to acute hypoxia, including under the influence of indralin, is associated with the release of adrenaline and noradrenaline. Catecholamines realize their effect through beta-adrenoceptors on lymphocytes. A close correlation was established between the dose and radioprotective effect of indralin and the growth of SDH lymphocyte activity (r = 0.99 p &lt; 0.01). Extrapolation of expected radioprotective effect of indralin per person was carried out according to the formula: DRF = 0.3988e0.009181x[r = 0.995 (0.78–0.9999) p &lt; 0.01], where x is the activity of SDH. DRF of indralin (100 mg intramuscularly) for humans was 1.6. When orally administered indralin at a dose of 450 mg DRF was 1.3.

Population balance modelling captures host cell protein dynamics in CHO cell cultures.

Monoclonal antibodies (mAbs) have been extensively studied for their wide therapeutic and research applications. Increases in mAb titre has been achieved mainly by cell culture media/feed improvement and cell line engineering to increase cell density and specific mAb productivity. However, this improvement has shifted the bottleneck to downstream purification steps. The higher accumulation of the main cell-derived impurities, host cell proteins (HCPs), in the supernatant can negatively affect product integrity and immunogenicity in addition to increasing the cost of capture and polishing steps. Mathematical modelling of bioprocess dynamics is a valuable tool to improve industrial production at fast rate and low cost. Herein, a single stage volume-based population balance model (PBM) has been built to capture Chinese hamster ovary (CHO) cell behaviour in fed-batch bioreactors. Using cell volume as the internal variable, the model captures the dynamics of mAb and HCP accumulation extracellularly under physiological and mild hypothermic culture conditions. Model-based analysis and orthogonal measurements of lactate dehydrogenase activity and double-stranded DNA concentration in the supernatant show that a significant proportion of HCPs found in the extracellular matrix is secreted by viable cells. The PBM then served as a platform for generating operating strategies that optimise antibody titre and increase cost-efficiency while minimising impurity levels.

Composite Bioinks With Mesoporous Bioactive Glasses-A Critical Evaluation of Results Obtained by In Vitro Experiments.

Besides osteoconductivity and a high degradation rate, mesoporous bioactive glasses (MBGs) are specific for their highly ordered channel structure and high specific surface area, making them suitable as drug and/or growth factor delivery systems. On the other hand, the mesoporous channel structure and MBG composition can have an effect on common cell evaluation assays, leading to inconclusive results. This effect is especially important when MBG is mixed in composite bioinks, together with cells. Additionally, the hydrogel component of the ink can influence the degradation of MBG, leading to different ion releases, which can additionally affect the analyses. Hence, our aim here was to show how the MBG structure and composition influence common cell viability and differentiation assays when calcium (Ca)- or magnesium (Mg)-containing glass is part of an alginate-based composite bioink. We suggested pre-labeling of cells with DiI prior to bioprinting and staining with calcein-AM to allow identification of metabolically active cells expressing signals in both green and red channels, allowing the use of fluorescence imaging for cell viability evaluations in the presence of high amounts (7 wt %) of MBGs. The release and uptake of ions during degradation of CaMBG and MgMBG were significantly changed by alginate in the composite bioinks, as confirmed by higher release and uptake from bulk glasses. Additionally, we detected a burst release of Mg2+ from composites only after 24 h of incubation. Furthermore, we demonstrated that released ions and the mesoporous channel structure affect the measurement of lactate dehydrogenase (LDH) and alkaline phosphatase activity (ALP) in bioprinted composite scaffolds. Measured LDH activity was significantly decreased in the presence of CaMBG. On the other hand, the presence of MgMBG induced increased signal measured for the ALP. Taken together, our findings show how composite bioinks containing MBGs can interfere with common analyses, obtaining misleading results.

Diagnosing subclinical facial eczema in cattle: does combining liver enzyme tests increase the accuracy of diagnosis?

ABSTRACT Aims To assess whether adding glutamate dehydrogenase (GDH) activity measurements to measurements of gamma-glutamyl transferase (GGT) activity appreciably increases the accuracy of diagnosis of subclinical facial eczema (FE) in cattle. Methods As part of a larger study on the impact of FE on productivity, GGT and GDH activities were measured in serum samples collected from 426 cattle from one dairy farm in the Taranaki region in April 2018. Bayesian latent class analysis was then used to estimate herd prevalence of subclinical FE as well as the specificity and sensitivity of the activity in serum of GGT or GDH alone, and of GGT and GDH activities combined, as diagnostic tests for subclinical FE. Results The latent class analysis estimated the true prevalence of subclinical FE in the study population as 47.5 (95% probability interval (PI) = 38.3–55.3)%. There was no evidence of any clinically relevant difference between GGT and GDH activities as predictors of subclinical FE; the difference between the areas under the receiver operating characteristic curves for the two measures was 0.005 (95% PI = –0.02 to –0.03). Using the two tests in parallel, with a threshold of 50 IU/L for GGT and 225 IU/L for GDH resulted in specificity and sensitivity of >95%, markedly increasing the accuracy of diagnosis of subclinical FE compared to using GGT or GDH alone at any threshold Conclusions and clinical relevance In this herd, combining the two tests resulted in a clinically relevant improvement in the accuracy of diagnosis of subclinical FE compared to using either test alone, which if used at the individual level will result in fewer cattle being assigned the wrong FE status. This will also apply at the herd level, with combined testing producing fewer false-positive herd test results than using one enzyme alone. This is particularly important for monitoring the efficacy of FE control measures when the expectation should be that the proportion of cattle with FE is very low.

Comparison between immunoassay technique and a manual laboratory method for the measurement of sorbitol dehydrogenase (SDH) activity for different types of diabetes in Nineveh governorate.

In this research ,the efficacy of the sorbitol dehydrogenase enzyme was compared with immunoassay technology and laboratory method for diabetic patients in Mosul city .Types of diabetic patient were divided in to four groups whose ages range from (8-62) years of both sexes (64 males and 56 females).The first group of patients who depend on their treatment on insulin injection only known as typ1 diabetes classified, and the second group was classified for patients who depend in their treatment on pills only to treat diabetes, which is known type 2 diabetes, while the third group of gestational diabetes was classified exclusively , in the four group of patients who depend on pills and injections in their treatment, the activity of (SDH) enzyme was estimated in the serum of healthy people without any mentioned disease, and they were considered a control group. The results showed a high significant in glucose concentration in serum of patient diabetes when compared with control group, also it was observed that there was a significant increase in the activity of the (SDH) enzyme in the serum of person with diabetes in the four group when compared with the control group which measured by the immunoassay technique and by the laboratory method. the results showed a significant increase in sorbitol dehydrogenase have been compared with the control group. The relationship was a study between (SDH) activity and glucose concentration of patient and control groups by finding the linear correlation coefficient. The positive significant correlation have found between the (SDH) activity of enzyme and the glucose concentration in control and patient groups.

Antibacterial Optimization of Highly Deformed Titanium Alloys for Spinal Implants.

The goal of the work was to develop materials dedicated to spine surgery that minimized the potential for infection originating from the transfer of bacteria during long surgeries. The bacteria form biofilms, causing implant loosening, pain and finally, a risk of paralysis for patients. Our strategy focused both on improvement of antibacterial properties against bacteria adhesion and on wear and corrosion resistance of tools for spine surgery. Further, a ~35% decrease in implant and tool dimensions was expected by introducing ultrahigh-strength titanium alloys for less-invasive surgeries. The tested materials, in the form of thin, multi-layered coatings, showed nanocrystalline microstructures. Performed direct-cytotoxicity studies (including lactate dehydrogenase activity measurement) showed that there was a low probability of adverse effects on surrounding SAOS-2 (Homo sapiens bone osteosarcoma) cells. The microbiological studies (e.g., ISO 22196 contact tests) showed that implanting Ag nanoparticles into Ti/TixN coatings inhibited the growth of E. coli and S. aureus cells and reduced their adhesion to the material surface. These findings suggest that Ag-nanoparticles present in implant coatings may potentially minimize infection risk and lower inherent stress.

Acute exposure to bis(2-ethylhexyl)phthalate disrupts calcium homeostasis, energy metabolism and induces oxidative stress in the testis of Danio rerio

Bis(2-ethylhexyl)phthalate (BEHP) negatively affects testicular functions in different animal species, disturbing reproductive physiology and male fertility. The present study investigated the in vitro acute effect of BEHP on the mechanism of action of ionic calcium (Ca2+) homeostasis and energy metabolism. In addition, the effect of BEHP on oxidative stress was studied in vitro and in vivo in the testis of Danio rerio (D. rerio). Testes were treated in vitro for 30 min with 1 μM BEHP for 45Ca2+ influx measurements. Testes were also incubated with 1 μM BEHP for 1 h (in vitro) or 12 h (in vivo) for the measurements of lactate content, 14C-deoxy-d-glucose uptake, lactate dehydrogenase (LDH) and gamma-glutamyl transpeptidase (GGT) activity, total reactive oxygen species (ROS) production and lipid peroxidation. In addition, the effect of BEHP (1 μM) on GGT, glutamic oxaloacetic transferase (GOT) and glutamic pyruvic transferase (GPT) activity in the liver was evaluated after in vivo treatment for 12 h. BEHP disturbs the Ca2+ balance in the testis when given acutely in vitro. BEHP stimulated Ca2+ influx occurs through L-type voltage-dependent Ca2+ channels (L-VDCC), transitory receptor potential vaniloid (TRPV1) channels, reverse-mode Na+/Ca2+ exchanger (NCX) activation and inhibition of sarco/endoplasmic reticulum Ca2+-ATPase (SERCA). BEHP affected energy metabolism in the testis by decreasing the lactate content and LDH activity. In vitro and in vivo acute effects of BEHP promoted oxidative stress by increasing ROS production, lipid peroxidation and GGT activity in the testis. Additionally, BEHP caused liver damage by increasing GPT activity.

Inhibitory effect of C-X-C motif chemokine ligand 14 on the osteogenic differentiation of human periodontal ligament cells through transforming growth factor-beta1

ObjectiveThis study aimed to determine the expression of chemokine (C-X-C motif) ligand 14 (CXCL14) in pulpal and periodontal cells in vivo and in vitro, and investigate function of CXCL14 and its underlying mechanism in the proliferation and osteogenic differentiation of human periodontal ligament (hPDL) cells. MethodsTo determine the expression level of CXCL14 in adult rat oral tissues and in hPDL cells after application of biophysical forces, RT-PCR, western blot, and histological analyses were performed. The role of CXCL14 in proliferation and osteogenic differentiation of PDL cells was evaluated by measuring dehydrogenase activity and Alizarin red S staining. ResultsStrong immunoreactivity against CXCL14 was observed in the PDL tissues and pulpal cells of rat molar, and attenuated apparently by orthodontic biophysical forces. As seen in rat molar, highly expressed CXCL14 was observed in human dental pulp and hPDL cells, and attenuated obviously by biophysical tensile force. CXCL14 expression in hPDL cells was increased in incubation time-dependent manner. Proliferation of hPDL cells was inhibited dramatically by small interfering (si) RNA against CXCL14. Furthermore, dexamethasone-induced osteogenic mineralization was inhibited by recombinant human (rh) CXCL14, and augmented by CXCL14 siRNA. rhCXCL14 increased transforming growth factor-beta1 (TGF- β1) in hPDL cells. Inhibition of the cell proliferation and osteogenic differentiation of hPDL cells by CXCL14 siRNA and rhCXCL14 were restored by rhTGF-β1 and SB431542, respectively. ConclusionThese results suggest that CXCL14 may play roles as a growth factor and a negative regulator of osteogenic differentiation by increasing TGF-β1 expression in hPDL cells.

Roflumilast, a phosphodiesterase-4 inhibitor, improves hyperoxia-induced lung injury via anti-inflammation

Roflumilast is an inhibitor of phosphodiesterase-4 (PDE4) and can suppress the hydrolysis of cAMP in inflammatory cells, conferring anti-inflammatory effects. This study aimed to investigate the protective effects of roflumilast on hyperoxia-induced acute lung injury (HALI) in a rat model. Male Sprague-Dawley rats were randomly assigned into: control group; HALI group; 2.5 mg/kg roflumilast group; and 5 mg/kg roflumilast group. Rats were pressurized to 250 kPa with pure oxygen to induce lung injury. In the roflumilast groups, rats were orally administered with roflumilast at 2.5 or 5 mg/kg once before hyperoxia exposure and once daily for two days after exposure. Rats were sacrificed 72 hours after hyperoxia exposure. The lung tissues were collected for the detection of lung water content, inflammatory cytokines and NF-κB/p-NF-κB protein expression, and the bronchoalveolar lavage fluid was harvested for the measurement of protein concentration and lactate dehydrogenase activity. Results showed roflumilast at different doses could significantly reduce lung edema, improve lung pathology and reduce the expression of inflammatory cytokines in the lung. The protective effects seemed to be related to the dose of roflumilast. Our study indicates roflumilast has the potential as a medication for the treatment of HALI.

Developing a method for measurement of dehydrogenase activity in biological wastewater treatment processes applied for toxic compounds degradation

Biological wastewater treatment processes are among the environmentally friendly techniques for degradation of organic compounds. They are also preferred to the physical and chemical processes which are due to the ability of biological processes to treat wide range of organic compounds with lower operational costs. However, biological processes are usually affected by variation in the inlet wastewater quality and quantity. In order to investigate the performance of the wastewater treatment plant, various parameters in case of effluent quality such as COD, BOD, TSS, TDS etc. are required to be measured. Microorganisms in bioreactors use various enzymes to degrade the organic contaminants. Higher toxic organic load on the biological process may lead to the deterioration of the process performance which is due to the reduction in microbial activity of the biomass. Dehydrogenase enzyme produced in biological processes could be used as an indicator for the biological wastewater treatment. Present study introduces a simple and modified method for evaluation of biological wastewater treatment process measuring dehydrogenase activity. In the present study, the effective parameters such as incubation time and types of solvent were investigated and the best procedure is developed for measuring the dehydrogenase activity in biological wastewater treatment process.

Dimethyl fumarate and monomethyl fumarate attenuate oxidative stress and mitochondrial alterations leading to oxiapoptophagy in 158N murine oligodendrocytes treated with 7β-hydroxycholesterol

Oxidative stress and mitochondrial dysfunction contribute to the pathogenesis of neurodegenerative diseases and favor lipid peroxidation, leading to increased levels of 7β-hydroxycholesterol (7β-OHC) which induces oxiapoptophagy (OXIdative stress, APOPTOsis, autoPHAGY). The cytoprotective effects of dimethylfumarate (DMF), used in the treatment of relapsing remitting multiple sclerosis and of monomethylfumarate (MMF), its main metabolite, were evaluated on murine oligodendrocytes 158 N exposed to 7β-OHC (50 μM, 24 h) with or without DMF or MMF (25 μM). The activity of 7β-OHC in the presence or absence DMF or MMF was evaluated on several parameters: cell adhesion; plasma membrane integrity measured with propidium iodide (PI), trypan blue and fluoresceine diacetate (FDA) assays; LDH activity; antioxidant enzyme activities (superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx)); generation of lipid peroxidation products (malondialdehyde (MDA), conjugated dienes (CDs)) and protein oxidation products (carbonylated proteins (CPs)); reactive oxygen species (ROS) overproduction conducted with DHE and DHR123. The effect on mitochondria was determined with complementary criteria: measurement of succinate dehydrogenase activity, evaluation of mitochondrial potential (ΔΨm) and mitochondrial superoxide anions (O2●−) production using DiOC6(3) and MitoSOX, respectively; quantification of mitochondrial mass with Mitotracker Red, and of cardiolipins and organic acids. The effects on mitochondrial and peroxisomal ultrastructure were determined by transmission electron microscopy. Intracellular sterol and fatty acid profiles were determined. Apoptosis and autophagy were characterized by staining with Hoechst 33,342, Giemsa and acridine orange, and with antibodies raised against caspase-3 and LC3. DMF and MMF attenuate 7β-OHC-induced cytotoxicity: cell growth inhibition; decreased cell viability; mitochondrial dysfunction (decrease of succinate dehydrogenase activity, loss of ΔΨm, increase of mitochondrial O2●− production, alteration of the tricarboxilic acid (TCA) cycle, and cardiolipins content); oxidative stress induction (ROS overproduction, alteration of GPx, CAT, and SOD activities, increased levels of MDA, CDs, and CPs); changes in fatty acid and cholesterol metabolism; and cell death induction (caspase-3 cleavage, activation of LC3-I in LC3-II). Ultrastructural alterations of mitochondria and peroxisomes were prevented. These results demonstrate that DMF and MMF prevent major dysfunctions associated with neurodegenerative diseases: oxidative stress, mitochondrial dysfunction, apoptosis and autophagy.