Can pumpkin save us of doxorubicin induced cardiotoxicity? / ¿Puede la calabaza salvarnos de la cardiotoxicidad inducida por doxorrubicina?

SUMMARY: Doxorubicin (DOX) is one of the drugs necessary for the treatment of the 10 most common types of cancer. The leading adverse effect limiting clinical use of DOX is cardiotoxicity. Given that literature data indicate a protective role of carotenoids in doxorubicin-induced toxicity, in our study we compared the cardioprotective effect of a mixture of pumpkin carotenoids and a commercially available antioxidant preparation. Animals were distributed in 8 groups (Control - S; NADES - N; Doxorubicin - Dox; Carotenoids - Car; CardiofortIN - CF; NADES-Doxorubicin - N-Dox; Carotenoids-Doxorubicin - Car-Dox; CardiofortIN-Doxorubicin - CF-Dox). Histological sections were stained with the hematoxylin-eosin (HE) and analyzed for the presence of myocardial damage by doxorubicin damage score (DDS). From the heart tissue homogenate were determined the intensity of lipid peroxidation and specific antioxidative enzyme activity (superoxide dismutase; catalase; glutathione S-transferase; glutathione peroxidase). In Car-DOX and CF-DOX groups, lipid peroxidation is significantly reduced compared to DOX group. Pretreatment of animals with carotenoids and in lesser extent with CardiofortIN led to higher antioxidative enzymes activity, compared to DOX group. Pretreated with carotenoids, only 50 % of animals had some degree of myocardial damage, and no animals had extensive damage. CardiofortIN pretreatment showed less protective effect. Pretreatment with carotenoid extract, reduced DDS significantly, so Car-DOX group has changes equivalent to mild myocardial damage. Although CardiofortIN pretreatment lowered DDS score values, animals still had moderate level of myocardium damage. This in vivo study and its findings indicate that carotenoids extracted from pumpkin may be a promising cardioprotective agent against doxorubicin induced cardiotoxicity, at least in part mediated through inhibition of DOX-induced oxidative stress.

La doxorrubicina (DOX) es uno de los fármacos necesarios para el tratamiento de los 10 tipos más comunes de cáncer. El principal efecto adverso que limita el uso clínico de DOX es la cardiotoxicidad. Debido a que los datos de la literatura indican un papel protector de los carotenoides en la toxicidad inducida por doxorrubicina, en nuestro estudio comparamos el efecto cardioprotector de una mezcla de carotenoides de calabaza y una preparación antioxidante disponible comercialmente. Los animales se distribuyeron en 8 grupos (Control - S; NADES - N; Doxorrubicina - Dox; Carotenoides - Car; CardiofortIN - CF; NADES-Doxorrubicina - N-Dox; Carotenoides-Doxorrubicina - Car-Dox; CardiofortIN- Doxorrubicina - CF-Dox). Las secciones histológicas se tiñeron con hematoxilina-eosina (HE) y se analizaron para detectar la presencia de daño miocárdico mediante la puntuación de daño por doxorrubicina (DDS). A partir del homogeneizado de tejido cardíaco se determinó la intensidad de la peroxidación lipídica y la actividad enzimática antioxidante específica (superóxido dismutasa, catalasa, glutatión S-transferasa, glutatión peroxidasa). En los grupos Car-DOX y CF-DOX, la peroxidación lipídica se redujo significativamente en comparación con el grupo DOX. El pre tratamiento de los animales con carotenoides y, en menor medida, con CardiofortlN condujo a una mayor actividad de las enzimas antioxidantes, en comparación con el grupo DOX. Al ser pre tratados con carotenoides, solo el 50 % de los animales tenían algún grado de daño miocárdico y ningún animal tenía daño extenso. El pre tratamiento con CardiofortIN mostró un efecto protector menor. El pre tratamiento con extracto de carotenoides redujo significativamente el DDS, por lo que el grupo Car-DOX mostró cambios equivalentes a un daño miocárdico leve. Aunque el pre tratamiento con CardiofortIN redujo los valores de la puntuación DDS, los animales aún tenían un nivel moderado de daño al miocardio. Este estudio in vivo y sus hallazgos indican que los carotenoides extraídos de la calabaza pueden ser un agente cardioprotector prometedor contra la cardiotoxicidad inducida por doxorrubicina, al menos en parte mediada por la inhibición del estrés oxidativo inducido por DOX.

Use of antioxidants and time of cold storage: effects over viability parameters and enzymatic levels in semen of rainbow trout (Oncorhynchus mykiss, Walbaum, 1792) / Uso de antioxidantes e tempo de armazenamento a frio: efeitos sobre parâmetros de viabilidade e níveis enzimáticos no sêmen da truta arco-íris (Oncorhynchus mykiss, Walbaum, 1792)

The cold storage of milt implies potentials alterations in its quality because the storage generates as main process, free radicals that produce spermatozoa membrane lipids damage with the consequent motility and fertilising capacity disruptions. To decrease the damage generated by free radicals the cells have antioxidant defences (proteins, enzymes, and low molecular weight substances). The objective of the present study evaluated the time storage effect and different antioxidants prepared in spermatic diluents on sperm viability of O. mykiss milt stored at 4°C. The two-way ANOVA denoted that the time storage and antioxidant influence have significant effects separated or combined on viability parameters (sperm motility and viability, proteins concentrations and superoxide dismutase enzymatic activity in seminal plasma). In contrast, only the storage time affected the fertilising capacity and catalase enzymatic activity in seminal plasma. The resulting analysis can conclude that the antioxidant presence improves the viability of cold stored milt, especially the transport conditions and the antioxidants allow the fecundity despite motility decrease.

O armazenamento a frio de leite implica potenciais alterações em sua qualidade, pois gera como processo principal radicais livres que provocam danos aos lipídios da membrana dos espermatozoides, com as consequentes alterações na motilidade e na capacidade de fertilização. Para diminuir os danos causados pelos radicais livres, as células têm defesas antioxidantes (proteínas, enzimas e substâncias de baixo peso molecular). O presente estudo avaliou o efeito do tempo de armazenamento e diferentes antioxidantes preparados em diluentes espermáticos no armazenamento de viabilidade de O. mykiss milt a 4°C. A ANOVA de duas vias denotou que o armazenamento no tempo e a influência antioxidante têm efeitos significativos separados ou combinados nos parâmetros de viabilidade (motilidade espermática, viabilidade espermática, concentrações de proteínas e atividade enzimática da superóxido dismutase no plasma seminal), enquanto apenas o tempo de armazenamento afetou a capacidade de fertilização e atividade enzimática da catalase no plasma seminal. A análise resultante pode concluir que a presença de antioxidante melhora a viabilidade do leite frio, especialmente as condições de transporte, e os antioxidantes permitem a fecundidade apesar da diminuição da motilidade.

The effects of metformin, acetylsalicylic acid and ibuprofen on telomerase enzyme activity: inhibitory effect of ibuprofen

Abstract Telomerase enzyme is necessary for the elongation of telomeres while telomerase being critical for aging and cancer. Metformin, ibuprofen, and acetylsalicylic acid used in this research are drugs that millions of people already use and that many are likely to use in future. In this study, the effects of these drugs on telomerase activity of Mus musculus swiss albino mice in liver tissue were investigated and the telomerase activity was measured with a PCR-ELISA based kit. In the study a possible connection between telomerase enzyme activity and activities of antioxidant enzymes was also investigated by determining the activity of superoxide dismutase (SOD) and catalase enzymes. The data obtained show that metformin slightly decreased telomerase enzyme activity in low dose application; however, this change was not statistically significant. In ibuprofen application, there was a significant inhibitory effect when high doses were used; whereas, there was a slight inhibitory effect at low doses. In acetylsalicylic acid application, a slight activator effect was detected; it was not statistically significant, though. Metformin was observed to increase catalase and SOD activities in general while low and high doses of acetyl salicylic acid showed different effects. In addition, ibuprofen caused a statistically significant increase in liver SOD values. It is important to note that this study demonstrated a significant inhibitory effect of ibuprofen on telomerase enzyme activity in animal models..

Multiple metals and agricultural use affects oxidative stress biomarkers in freshwater Aegla crabs / Presença de múltiplos metais e agricultura afetam biomarcadores de estresse oxidativo em caranguejos de água doce (Aegla)

Metals and agrochemicals are among the main aquatic contaminants, being able to trigger oxidative stress in exposed organisms. The objective of this work was to evaluate the correlation between the level of oxidative stress biomarkers in Aegla crabs (Crustacea, Anomura) with (i) the set of metals present in the streams sediment and (ii) with land uses of three hydrographic basins. The study was carried out in streams (≤ 2nd order) of hydrographic basins in southern Brazil (Basins of Rio Suzana, Rio Ligeirinho-Leãozinho and Rio Dourado). In these streams were quantified the land uses and Cu, Cr, Cd, Fe, Mn and Zn concentrations in the sediment. The enzymes Catalase (CAT) and Glutathione Reductase (GR), as well as the level of membrane lipid peroxidation (TBARS), were analyzed in adult females. The PCA analysis showed that the distribution of metals was different between the basins. Cd, Cr and Fe were correlated positively with CAT and negatively with TBARS and GR. The Dourado basin had the lowest concentrations of these three metals and the highest levels of TBARS. However, in Dourado basin there is predominance of agriculture land use, and TBARS was positively correlated with agricultural land use. Besides in Dourado basin, GR activity was higher than in the others basins, indicating a compensatory response in relation to CAT inhibition. The basins of Suzana and Ligeirinho-Leãozinho rivers had lower TBARS values, which may be due to the induction of CAT in response to metals accumulated in sediment. In summary, this work indicates that in the basins with a higher concentration of toxic metals there is an adaptive response of CAT induction, which reduces TBARS in Aegla. On the other hand, in the basin with lower metallic contamination, TBARS occurrence was primarily influenced by agricultural land use.

Os metais e agroquímicos estão entre os principais contaminantes aquáticos, podendo desencadear estresse oxidativo em organismos expostos. O objetivo deste trabalho foi avaliar uma possível correlação entre o nível de biomarcadores de estresse oxidativo em Aegla (Crustacea, Anomura) com (i) o conjunto de metais presentes no sedimento e (ii) com os usos da terra, em três bacias hidrográficas distintas. O estudo foi realizado em riachos(≤ 2ª ordem) de bacias hidrográficas do Sul do Brasil (Bacias do Rio Suzana, do Rio Ligeirinho-Leãozinho e do Rio Dourado), as quais foram caracterizadas em função do percentual de usos da terra e do nível de Cu, Cr, Cd, Fe, Mn e Zn no sedimento. As enzimas Catalase (CAT) e Glutationa Redutase (GR), bem como o nível de peroxidação lipídicadas membranas (TBARS), foram analisadas em fêmeas adultas. Uma análise de PCA mostrou que a distribuição de metais foi distinta entre as bacias. Cd, Cr e Fe no sedimento correlacionaram positivamente com a CAT e negativamente com TBARS e GR. Entretanto, a bacia do Dourado apresentou os menores níveis destes três metais e os maiores níveis de TBARS, o que pode ser justificado pelo predomínio da agricultura nesta bacia, já que o TBARS correlacionou positivamente com o percentual de uso agrícola. Nesta bacia, a atividade da GR foi mais alta do que nas outras, indicando uma resposta compensatória em relação a inibição da CAT. As bacias do rio Suzana e rio Ligeirinho-Leãozinho apresentaram valores menores de TBARS, o que pode decorrer da indução da CAT em função dos metais acumulados no sedimento. Em síntese, este trabalho indica que nas bacias com maior concentração de metais tóxicos ocorre uma resposta adaptativa de indução da CAT, o que reduz os níveis de TBARS em Aegla. Por outro lado, na bacia com menor contaminação metálica os níveis de TBARS foram primariamente influenciados pelo uso agrícola.

Preparation and catalytic properties of catalase-inorganic hybrid nanoflowers / 生物工程学报

Catalase is widely used in the food, medical, and textile industries. It possesses exceptional properties including high catalytic efficiency, high specificity, and environmental friendliness. Free catalase cannot be recycled and reused in industry, resulting in a costly industrial biotransformation process if catalase is used as a core ingredient. Developing a simple, mild, cost-effective, and environmentally friendly approach to immobilize catalase is anticipated to improve its utilization efficiency and enzymatic performance. In this study, the catalase KatA derived from Bacillus subtilis 168 was expressed in Escherichia coli. Following separation and purification, the purified enzyme was prepared as an immobilized enzyme in the form of enzyme-inorganic hybrid nanoflowers, and the enzymatic properties were investigated. The results indicated that the purified KatA was obtained through a three-step procedure that included ethanol precipitation, DEAE anion exchange chromatography, and hydrophobic chromatography. Then, by optimizing the process parameters, a novel KatA/Ca3(PO4)2 hybrid nanoflower was developed. The optimum reaction temperature of the free KatA was determined to be 35 ℃, the optimum reaction temperature of KatA/Ca3(PO4)2 hybrid nanoflowers was 30-35 ℃, and the optimum reaction pH of both was 11.0. The free KatA and KatA/Ca3(PO4)2 hybrid nanoflowers exhibited excellent stability at pH 4.0-11.0 and 25-50 ℃. The KatA/Ca3(PO4)2 hybrid nanoflowers demonstrated increased storage stability than that of the free KatA, maintaining 82% of the original enzymatic activity after 14 d of storage at 4 ℃, whereas the free KatA has only 50% of the original enzymatic activity. In addition, after 5 catalytic reactions, the nanoflower still maintained 55% of its initial enzymatic activity, indicating that it has good operational stability. The Km of the free KatA to the substrate hydrogen peroxide was (8.80±0.42) mmol/L, and the kcat/Km was (13 151.53± 299.19) L/(mmol·s). The Km of the KatA/Ca3(PO4)2 hybrid nanoflowers was (32.75±2.96) mmol/L, and the kcat/Km was (4 550.67±107.51) L/(mmol·s). Compared to the free KatA, the affinity of KatA/Ca3(PO4)2 hybrid nanoflowers to the substrate hydrogen peroxide was decreased, and the catalytic efficiency was also decreased. In summary, this study developed KatA/Ca3(PO4)2 hybrid nanoflowers using Ca2+ as a self-assembly inducer, which enhanced the enzymatic properties and will facilitate the environmentally friendly preparation and widespread application of immobilized catalase.

Prevalence and effect of intramammary infection due to coagulase-negative staphylococcal (CNS) species in somatic cell counts in milk from Holstein dairy cows in Boyaca, Colombia / Prevalencia y efecto de la infección intramamaria debida a especies de estafilococos coagulasa negativo (ECN) en el conteo de células somáticas en leche de vacas Holstein en Boyacá, Colombia

ABSTRACT Mastitis is one of the most important illnesses in specialized dairy herds worldwide due to the effects on production and animal health. The types caused by CNS has a special importance in a production where the main pathogens are controlled. The objective of the present work is to determine the prevalence of CNS in a dairy herd in Boyaca and also quantify the effects of every species of CNS in SCC. 40 cows were selected and sampled during 6 months, CMT was performed, and results from 1 to trace were sampled. The routine bacteriological test was also performed for CNS identification, and the isolating of CNS was performed through rpoB gene identification and through the type of strain using the pulse gel electrophoresis procedure. Out of 960 samples, 619 were positive for CNS growth. The most prevalent species were Staphylococcus epidermidis, S. chromogenes, S. sciuri, S. simulaans, S. haemolyticus and S. capitis. The results that were found here are similar to the results observed in different parts of the world, which confirms that they are pathogens that must be constantly evaluated because they can go unnoticed in routine controls, especially in those farms where major pathogens are not a serious problem. The results determined in this study demonstrate that CNS generates a slight increase in somatic cells.

RESUMEN La mastitis es una de las enfermedades más importantes en los rebaños lecheros especializados alrededor de todo el mundo debido a los efectos sobre la producción y la salud animal. Los tipos ocasionados por estafilococos coagualasa negativo (ECN) tienen una importancia especial en una producción en la que los principales patógenos están controlados. El objetivo del presente trabajo es determinar la prevalência del ECN en un hato lechero en Boyacá y cuantificar los efectos de cada especie de ECN en el conteo de células somáticas (CCS). Se seleccionaron 40 vacas y se tomaron muestras durante 6 meses, se realizó california mastitis test (CMT) y se tomaron muestras de los resultados desde 1 hasta donde hubo trazas. También se realizó la prueba bacteriológica de rutina para la identificación del ECN y el aislamiento del ECN se realizó mediante la identificación del gen rpoB y del tipo de cepa, usando el procedimiento de electroforesis en gel de pulso. De 960 muestras, 619 fueron positivas para el crecimiento del ECN. Las especies más prevalentes fueron Staphylococcus epidermidis, S. chromogenes, S. sciuri, S. simulans, S. haemolyticus y S. capitis. Los resultados encontrados aquí son similares a resultados en diferentes partes del mundo, lo que confirma que son patógenos que deben ser evaluados constantemente porque pueden pasar desapercibidos en los controles de rutina, especialmente en aquellas fincas donde los patógenos mayores no son un problema grave. Los resultados determinados en este estudio demuestran que el SNC genera un ligero aumento de células somáticas.

The effects of rapamycin and 3-methyladenine on nitric oxide synthase enzymes in cisplatin-induced neurotoxicity / Efectos de la rapamicina y la 3-metiladenina sobre la enzima óxido nítrico sintasa en la neurotoxicidad inducida por cisplatino

SUMMARY: The aim of this study was to determine the relationship of autophagy-enhancing rapamycin (RAPA) and autophagy- inhibitor 3-methyladenine (3-MA) with Nitric oxide synthases (NOS) in Cisplatin (CIS)-induced neurotoxicity in rats. Rats were divided into 4 groups (n=10): Control was applied saline, CIS (a single dose of 8mg/kg intraperitoneal (i.p.) on 7th day of experiment), RAPA+CIS (2 mg/kg/i.p. RAPA per day and 8 mg/kg/i.p. CIS on 7th day), 3-MA+CIS (15 mg/kg/i.p. 3-MA per day and 8 mg/kg/i.p. CIS on 7th day). Rats were sacrificed under anesthesia. Brain tissues were evaluated histopathologically. eNOS, Inos, nNOS and MAP 2 immunostaining were performed to determine the expression levels of these proteins among groups. Superoxide dismutase (SOD), Catalase (CAT), Malondialdehyde (MDA) and Interleukin IL-6 levels in brain tissue and serum nitric oxide (NO) level were measured by ELISA assay. In histopathological evaluation, neurodegeneration was seen in the CIS group. There was an increase in eNOS, iNOS and nNOS immunostaining in CIS group. While MAP2 immunostaining of the CIS group decreased. There was a decrease in SOD and CAT levels of brain tissue in CIS group. However, there was an increase in MDA, IL-6 and NO levels of brain tissue in CIS group. We found that antioxidant capacity increase while, inflammation and nitric oxide levels decreased in the RAPA-treated group. 3-MA does not have a significant effect. We suggest that CIS-induced neurotoxicity is more effective than Rapa 3-MA and may also be linked to NOS enzymes.

RESUMEN: El objetivo de este estudio fue determinar la relación de la rapamicina potenciadora de la autofagia (RAPA) y el inhibidor de la autofagia 3-metiladenina (3-MA) con óxido nítrico sintasas (NOS) en la neurotoxicidad inducida por cisplatino (CIS) en ratas. Las ratas se dividieron en 4 grupos (n = 10): grupo control se aplicó solución salina, CIS (una dosis única de 8 mg / kg intraperitoneal (ip) el día 7 del experimento), RAPA + CIS (2 mg / kg / ipRAPA por día y 8 mg / kg / ip CIS el día 7), 3-MA + CIS (15 mg / kg / ip 3-MA por día y 8 mg / kg / ip CIS el día 7). Las ratas se sacrificaron bajo anestesia y los tejidos cerebrales fueron analizados histopatológicamente. Se realizaron inmunotinciones con eNOS, Inos, nNOS y MAP 2 para determinar los niveles de expre- sión de estas proteínas entre los grupos. Se midieron los niveles de superóxido dismutasa (SOD), catalasa (CAT), malondialdehído (MDA) e interleucina IL-6 en el tejido cerebral y el nivel de óxido nítrico (NO) en suero mediante ensayo ELISA. En la evaluación histopatológica, se observó neurodegeneración en el grupo CIS. Hubo un aumento en la inmunotinción de eNOS, iNOS y nNOS en el grupo CIS. Mientras que la inmunotinción de MAP2 del grupo CIS disminuyó. Hubo una disminución en los niveles de SOD y CAT del tejido cerebral en el grupo CIS, sin embargo, hubo un aumento en los niveles de MDA, IL-6 y NO en el tejido cerebral en el grupo CIS. Observamos que la capacidad antioxidante aumentó, mientras que la inflamación y los niveles de óxido nítrico disminuyeron en el grupo tratado con RAPA. 3-MA no tiene un efecto significativo. Sugerimos que la neurotoxicidad inducida por CIS es más eficaz que Rapa 3-MA y también puede estar relacio- nada con las enzimas NOS.

Treino de Força Reduz Stress Oxidativo Cardíaco e Renal em Ratos com Hipertensão Renovascular / Strength Training Reduces Cardiac and Renal Oxidative Stress in Rats with Renovascular Hypertension

Resumo Fundamento O treino de força tem efeitos benéficos em doenças renais, além de ajudar a melhorar a defesa antioxidante em animais saudáveis. Objetivo Verificar se o treino de força reduz o dano oxidativo ao coração e rim contralateral para cirurgia de indução de hipertensão renovascular, bem como avaliar as alterações na atividade das enzimas antioxidantes endógenas superóxido dismutase (SOD), catalase (CAT) e glutationa peroxidase (GPx). Métodos Dezoito ratos machos foram divididos em três grupos (n=6/grupo): placebo, hipertenso e hipertenso treinado. Os animais foram induzidos a hipertensão renovascular através da ligação da artéria renal esquerda. O treino de força foi iniciado quatro semanas após a indução da hipertensão renovascular, teve 12 semanas de duração e foi realizada a 70% de 1RM. Depois do período de treino, os animais foram submetidos a eutanásia e o rim esquerdo e o coração foram retirados para realizar a quantificação de peróxidos de hidrogênio, malondialdeído e grupos sulfidrílicos, que são marcadores de danos oxidativos. Além disso, foram medidas as atividades das enzimas antioxidantes superóxido dismutase, catalase e glutationa peroxidase. O nível de significância adotado foi de 5% (p < 0,05). Resultados Depois do treino de força, houve redução de danos oxidativos a lipídios e proteínas, como pode-se observar pela redução de peróxidos de hidrogênio e níveis sulfidrílicos totais, respectivamente. Além disso, houve um aumento nas atividades das enzimas antioxidantes superóxido dismutase, catalase e glutationa peroxidase. Conclusão O treino de força tem o potencial de reduzir danos oxidativos, aumentando a atividades de enzimas antioxidantes. (Arq Bras Cardiol. 2021; 116(1):4-11)

Abstract Background Strength training has beneficial effects on kidney disease, in addition to helping improve antioxidant defenses in healthy animals. Objective To verify if strength training reduces oxidative damage to the heart and contralateral kidney caused by the renovascular hypertension induction surgery, as well as to evaluate alterations in the activity of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) endogenous antioxidant enzymes. Methods Eighteen male rats were divided into three groups (n=6/group): sham, hypertensive, and trained hypertensive. The animals were induced to renovascular hypertension through left renal artery ligation. Strength training was initiated four weeks after the induction of renovascular hypertension, continued for a 12-weeks period, and was performed at 70% of 1RM. After the training period, the animals were euthanized and the right kidney and heart were removed for quantitation of hydroperoxides, malondialdehyde and sulfhydryl groups, which are markers of oxidative damage. In addition, the activity of SOD, CAT, and GPx antioxidant enzymes was also measured. The adopted significance level was 5% (p < 0.05). Results After strength training, a reduction in oxidative damage to lipids and proteins was observed, as could be seen by reducing hydroperoxides and total sulfhydryl levels, respectively. Furthermore, an increased activity of superoxide dismutase, catalase, and glutathione peroxidase antioxidant enzymes was observed. Conclusion Strength training is able to potentially reduce oxidative damage by increasing the activity of antioxidant enzymes. (Arq Bras Cardiol. 2021; 116(1):4-11)

Effect of Astaxanthin on Antioxidant Enzyme Activities in Suspended Leukocyte-Depleted Red Blood Cells Stored for Transfusion / 中国实验血液学杂志

OBJECTIVE@#To observe the effect of astaxanthin (ASTA) on the activities of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) in suspended leukocyte-depleted red blood cells stored for transfusion.@*METHODS@#The suspended leukocyte-depleted red blood cells were randomly divided into group A, B, C and D. The ASTA was added into preservation solution of suspended leukocyte-depleted red blood cells of group B, C and D with the final concentration 5, 10 and 20 μmol/L, respectively, while DMSO was added into cells of group A in the same volume. After 7, 14, 28 and 42 days of storage, the reactive oxygen species (ROS) content in red blood cells was detected by fluorescence microplate reader, malondialdehyde (MDA) content was detected by thiobarbituric acid (TBA) method, activity of SOD was detected by xanthine oxidase method, the activity of CAT was detected by visible light method, and activity of GSH-Px was detected by colorimetry.@*RESULTS@#After 7, 14, 28 and 42 days of storage, the contents of ROS and MDA in suspended red blood cells of group B, C and D were significantly lower(P<0.05), while the activities of SOD and GSH-Px were higher than those of group A(P<0.05); and CAT activity in cells treated by ASTA was significantly higher at 28 and 42 days of storage in comparison with that of group A(P<0.05). There were positive correlations between the ROS, MDA content in suspended red blood cells of group A, B, C, D and storage time(P<0.01), while negative correlation between SOD, CAT, GSH-Px activity and storage time(P<0.01).@*CONCLUSION@#ASTA can decrease the oxidative stress level and peroxide damage degree by increasing the antioxidant enzyme activities in suspended leukocyte-depleted red blood cells during storage.

Effects of drought stress on physiological and biochemical and chemical components of Cinnamomum cassia seedlings / 中国中药杂志

Six month old Cinnamomum cassia seedlings were used to simulate drought stress with polyethylene glycol(PEG 6000). The physiological indicators(osmotic substances, antioxidant enzymes, etc.) and chemical components of seedlings under different drought levels and the correlation between the two were studied. The results showed that the chlorophyll content and relative water content decreased gradually with the increase of PGE 6000(0, 5%, 10%, 15%) concentration and time(3, 5, 7 d), while the soluble protein content, soluble sugar content and catalase(CAT) activity increased, but the rising rate slowed down with the time. The activities of peroxidase(POD), superoxide dismutase(SOD), malondialdehyde(MDA) and proline content increased at first and then decreased. The content of coumarin, cinnamaldehyde, cinnamic acid and dimethoxycinnamaldehyde decreased, while the content of cinnamyl alcohol continued to increase.Under drought stress, the fluorescence signals of reactive oxygen species and no contents in roots of C. cassia seedlings were significantly stronger than those of the control.Further correlation analysis showed that coumarin content, di-methoxycinnamaldehyde content and osmoregulation substance content were significantly negatively correlated(P<0.05), cinnamic acid content was significantly negatively correlated with POD and SOD activities(P<0.01).It was found that C. cassia seedlings showed a certain degree of drought tolerance under short-term or mild drought stress, but if the drought exceeded a certain degree, the physiological metabolism of the seedlings would be unbalanced.

Evaluation of Multidrug Resistant Loop-mediated Isothermal Amplification Assay for Detecting the Drug Resistance of / 生物医学与环境科学（英文）

Objective@#To evaluate multidrug resistant loop-mediated isothermal amplification (MDR-LAMP) assay for the early diagnosis of multidrug-resistant tuberculosis and to compare the mutation patterns associated with the @*Methods@#MDR-LAMP assay was evaluated using 100 @*Results@#The sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of MDR-LAMP were 85.5%, 93.6%, 96.7%, and 74.4% for the detection of resistance to isoniazid and rifampicin, respectively, and 80.5%, 92.3%, 98.6%, and 41.4% for the detection of @*Conclusion@#MDR-LAMP is a rapid and accessible assay for the laboratory identification of rifampicin and isoniazid resistance of

Fusion expression with catalase improves the stability of multicopper oxidase and its efficiency in degrading biogenic amines / 生物工程学报

Some enzymes belonging to the multicopper oxidase (MCO) family can degrade the hazardous biogenic amine (BA) present in food. However, the oxidation of MCO in the process of degrading BAs may reduce its activity and stability, resulting in decreased catalytic efficiency. In this work, an MCO from Lactobacillus fermentum (MCOF) was fused with a Bacillus subtilis catalase (CAT) using different strategies and the fusion enzymes were respectively expressed in Escherichia coli BL21(DE3). The tolerance of eight fused MCOFs to H2O2 increased by 51%-68%, and the stability of CAT&MCOF increased by 17%, compared to the wild type MCOF. Using 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) as a substrate, the substrate affinity (Km), the catalytic efficiency (kcat/Km) and the molar specific activity of CAT&MCOF increased by 1.0-fold, 1.7-fold and 1.2-fold than those of MCOF, respectively. The stability of CAT&MCOF under acidic conditions (pH 2.5-4.5) and moderate temperatures (35-55 °C) also improved. Moreover, the degradation rates of putrescine, cadaverine and histamine catalyzed by CAT&MCOF reached 31.7%, 36.0% and 57.8%, respectively, which increased by 132.5%, 45.7% and 38.9% compared to that of MCOF. The improvement on the stability and catalytic efficiency of MCOF by fusion expression with CAT provides a good example for improving the applicability of enzymes through molecular modifications.

Antioxidant activity of selenium on bisphenol-induced apoptosis and testicular toxicity of Albino rats / Actividad antioxidante el selenio sobre la apoptosis inducid por bisfenol y toxicidad testicular en ratas albinas

SUMMARY: Bisphenol A (BPA) is an industrial chemical widely used to make polycarbonate plastics for packaging and epoxy resins. This study sought to examine how selenium (Se) affects BPA toxicity in terms of albino rats' histological structure, antioxidant enzymes and reproductive organs (seminiferous tubules). Twenty-four adult male rats were divided into four experimental groups: Group 1: Control; Group 2: Orally administered BPA; Group 3: Orally administered sodium selenite; Group 4: Treated daily with BPA followed by selenium (Se). All experiment done for 4 weeks. BPA exposure caused changes in the testicular histological structure, which consists apoptosis, and led to changes in several biochemical markers: Malondialdehyde, catalase, superoxide dismutase, and glutathione peroxidase. However, these BPA side effects may be ameliorated in rats treated with BPA-plus-Se. These protective effects of Se may attributable to its ability to remove potentially damaging oxidizing agents in living organisms. The results may confirm that Se countered the oxidant effects and increased the BPA-induced stress response in rats. So, Se promotes the healthy growth and development of mammals by protecting them from oxidative stress. As human are greatly exposed to BPA and it can accumulate in tissues, there is concern about human reproductive functions particularly for occupational workers exposed usually to greater levels of BPA. Thus, the use of BPA in multiple industries must be restricted and the inaccurate usage of plastic containers should be avoided to decrease the health hazards. Administration of Se may protect against the adverse effects of BPA on reproductive functions and structures.

RESUMEN: El bisfenol A (BPA) es un químico industrial ampliamente utilizado para fabricar plásticos de policarbonato para envases y resinas epoxi. Este estudio examinó el efecto de selenio (Se) en la toxicidad del BPA en términos de la estructura histológica, enzimas antioxidantes y los órganos reproductivos (túbulos seminíferos) de ratas albinas. Se dividieron veinticuatro ratas macho adultas en cuatro grupos experimentales: Grupo 1: control; Grupo 2: BPA administrado por vía oral; Grupo 3: BPA administrado por vía oral para; Grupo 4: tratado diariamente con BPA seguido de selenio (Se). El experimento se realizó durante cuatro semanas y se observó que la exposición al BPA provocó cambios en la estructura histológica testicular, incluyendo apoptosis, y alteraciones en varios marcadores bioquímicos:malondialdehído, catalasa, superóxido dismutasa y glutatión peroxidasa. Sin embargo, estos efectos secundarios del BPA pueden mejorar en ratas tratadas con BPA-plus-Se. Estos efectos protectores del Se pueden ser atribuidos a la capacidad de eliminar agentes oxidantes potencialmente dañinos en organismos vivos. Los resultados indicaron que se contrarrestaron los efectos oxidantes y aumentó la respuesta al estrés inducido por BPA en ratas, y favorece el crecimiento y desarrollo en los mamíferos al protegerlos del estrés oxidativo. Debido a la exposición al BPA en el ser humano, se puede acumular en los tejidos, por lo que existe una preocupación por el daño a las funciones reproductivas en particular de los trabajadores que generalmente están expuestos a niveles más altos de BPA. Por lo tanto, se debe restringir el uso de BPA en las industrias y evitar el uso incorrecto de envases de plástico para así disminuir los riesgos para la salud. La administración correcta de Se puede proteger contra los efectos adversos del BPA en las funciones y estructuras reproductivas.

Bacteriemia por Lactococcus garvieae: primer caso en Colombia / Lactococcus garvieae bloodstream infections: first case in Colombia

Lactococcus garvieae es una causa inusual de bacteriemia. No existen guías diagnósticas y terapéuticas actuales disponibles para tratar las infecciones causadas por estos organismos. Con base en un informe de caso, proporcionamos una revisión de la literatura sobre bacteriemia causada por L. garvieae y destacamos los desafíos de diagnóstico y tratamiento de estas infecciones y las implicaciones para el manejo. Comunicamos el primer caso de bacteriemia por Lactococcus garvieae en Colombia.

Lactococcus garvieae is an unusual cause of bloodstreams infections. Currently, there are no diagnostic and therapeutic guides available to treat infections caused by these organisms. Based on a case report, we provide a literature review on bloodstreams caused by L. garvieae and highlighted the challenges for diagnose and treatment and direct implications for its management. We report the first case of bloodstream infections due to Lactococcus garvieae in Colombia.

Biomarcadores enzimáticos e histológicos em brânquias de Ucides cordatus (Linnaeus, 1763) (Crustacea, Brachyura, Ucididae) indicativos de impactos ambientais em uma região portuária do nordeste do Brasil / [Enzymatic and histological biomarkers in Ucides cordatus (Linnaeus, 1763) (Crustacea, Brachyura, Ucididae) gills indicative of environmental impacts in a port region of northeastern Brazil]

Objetivou-se neste estudo analisar biomarcadores histológicos e bioquímicos em brânquias de U. cordatus indicativos de impactos na Baía de São Marcos. Caranguejos foram coletados em quatro áreas na Baía de São Marcos: A1= Ilha dos Caranguejos (com baixo impacto); A2= Coqueiro, A3= Porto Grande, A4= Cajueiro (áreas potencialmente impactadas). Mediram-se os dados biométricos de cada exemplar de caranguejo. Amostras de brânquias foram submetidas à técnica histológica padrão e homogeneizadas em tampão fosfato, e o sobrenadante foi utilizado para análise das enzimas glutationa-S-transferase (GST) e catalase (CAT). A biometria indicou que os caranguejos de A1 são significativamente (P<0,05) maiores e mais pesados do que os caranguejos das áreas A2, A3 e A4. As alterações branquiais (rompimento das células pilastras, deformação do canal marginal, deslocamento da cutícula e necrose) foram significativamente (P˂0,05) mais frequentes em caranguejos de A2, A3 e A4 do que nos caranguejos de A1. As atividades enzimáticas da GST e CAT nos caranguejos apresentaram diferença significativa (P<0,05) entre as áreas de coletas, com padrão similar ao observado para as alterações branquiais. Os biomarcadores analisados mostraram que os caranguejos estão sob diferentes níveis de impactos (A4>A3>A2>A1) ao longo da Baía de São Marcos.(AU)

The objective of this study was to analyze histological and biochemical biomarkers in U. cordatus gills indicative of impacts in São Marcos Bay. Crabs were collected from four areas in São Marcos Bay: A1=Ilha dos Caranguejos (with low impact); A2=Coqueiro, A3=Porto Grande, A4=Cajueiro (potentially impacted areas). The biometric data of each specimen was measured. Gill samples were submitted to standard histological technique and homogenized in phosphate buffer, and the supernatant was used for analysis of glutathione S-transferase (GST) and catalase (CAT) enzyme activity. Biometric data indicated that crabs found in A1 are significantly (P<0.05) larger and heavier than crabs found in A2, A3 and A4 areas. Gill alterations (rupture of pilaster cells, Dilation of the marginal channel, Cuticle Rupture and necrosis) were significantly (P˂0.05) more frequent in the crabs in A2, A3 and A4 than crabs in A1. The enzymatic activities of GST and CAT showed significant difference (P<0.05) between the sampling areas, similar to that observed for gill alterations. The biomarkers analyzed showed that the crabs are under different impact levels (A4> A3> A2> A1) along the São Marcos Bay.(AU)

Stereological study and analysis of oxidative stress during renal aging in rats

Abstract Purpose To evaluate renal histological changes by stereology and morphometry and analyze the main markers of oxidative stress in rats undergoing natural aging. Methods Seventy two Wistar rats were divided into six groups of 12 rats each, which were euthanized at 3, 6, 9, 12, 18, and 24 months of age. Right kidney was stereologically and morphometrically analyzed to calculate the volumetric density (Vv[glom]), numerical density (Nv[glom]) and glomerular volume (Vol[glom]). Left kidney was used to determine the levels of nonprotein thiols, lipid peroxidation, and protein carbonylation, as well as the activities of superoxide-dismutase and catalase enzymes. Results Both Vv[glom] and Nv[glom] values showed gradual decreases between groups. Activity of superoxide-dismutase was elevated at 24 months of age, and the levels of nonprotein thiols were higher in older animals. Greater catalase activity and protein carbonylation were observed in animals between 6 and 12 months of age but lessened in older rats. Lipid peroxidation decreased in the older groups. Conclusions Morphometric and stereological analyses revealed a gradual decrease in the volume and density of renal glomeruli during aging, as well as kidney atrophy. These findings related to oxidative stress clarify many changes occurring in kidney tissues during senescence in rats.

Renal protective effects of vicenin-2 and scolymoside in a mouse model of sepsis

This study was initiated to determine whether 2 structurally related flavonoids found in Cyclopia subternata-vicenin-2 (VCN) and scolymoside (SCL)-could modulate renal functional damage in a mouse model of sepsis, and to elucidate the relevant underlying mechanisms. The potential of VCN and SCL treatment to reduce renal damage induced by cecal ligation and puncture (CLP) surgery in mice was measured via assessment of serum creatinine, blood urea nitrogen (BUN), lipid peroxidation, total glutathione, glutathione peroxidase activity, catalase activity, and superoxide dismutase activity. Treatment with either VCN or SCL resulted in elevated plasma levels of BUN and creatinine, and of protein in the urine of mice with CLP-induced renal damage. Moreover, both VCN and SCL inhibited nuclear factor κB activation and reduced the induction of nitric oxide synthase and excessive production of nitric acid. VCN and SCL treatment also reduced the plasma levels of interleukin-6, and tumor necrosis factor-α, reduced lethality due to CLP-induced sepsis, increased lipid peroxidation, and markedly enhanced the antioxidant defense system by restoring the levels of superoxide dismutase, glutathione peroxidase, and catalase in kidney tissues. The present results suggest that VCN and SCL protect mice from sepsis-triggered renal injury

Efeitos da vareniclina no tecido pulmonar em modelo animal / Effects of varenicline on lung tissue in the animal model

RESUMO Objetivo O objetivo deste estudo foi investigar os efeitos agudos e crônicos da vareniclina no tecido pulmonar em um estudo experimental. Métodos Um total de 34 ratos foi alocado aleatoriamente em grupos de estudo (vareniclina) e controle. Assim, os ratos foram divididos em dois grupos: (i) grupo controle e (ii) grupo vareniclina. A seguir, os ratos de cada grupo foram, por sua vez, subdivididos igualmente em agudos (C1; V1) e crônicos (C2; V2), e todos os ratos dos grupos agudos e crônicos foram sacrificados sob anestesia: no 45.º dia, para o grupo agudo [C1 (n=5) e V1 (n=12)], e no 90.º dia, para o grupo crônico [C2 (n=5) e V2 (n=12)], respectivamente. Em seguida, foram realizadas análises bioquímicas e histopatológicas. Resultados Trinta e quatro ratos completaram o estudo. Destes ratos, 24 estavam no grupo vareniclina e 10 no grupo controle. Na exposição crônica à vareniclina, os níveis de oxidante composto por malondialdeído (MDA) e mieloperoxidase (MPO) aumentaram, e os níveis de superóxido dismutase (SOD), catalase (CAT), glutationa (GSH) e glutationa peroxidase (GPx), nomeados como antioxidantes, diminuiram significativamente quando comparados com o grupo controle. Os níveis de MDA e MPO também foram significativamente mais elevados e os níveis de SOD, CAT, GPx e GSH foram significativamente mais baixos no grupo vareniclina crônico, quando comparado ao grupo vareniclina agudo. Estes achados também foram confirmados por observações histopatológicas. Conclusões Este é o primeiro estudo que avaliou os efeitos pulmonares da vareniclina experimentalmente em um modelo animal. Observamos que o tratamento crônico da vareniclina causa inflamação e lesão pulmonar.

ABSTRACT Objective This study aimed to investigate acute and chronic effects of varenicline on lung tissue in an experimental study. Methods A total of 34 rats were randomly allocated into study (varenicline) and control groups. The rats were divided into two groups (i) control group, (ii) varenicline group. Then, the rats in the each group were sub-divided equally in turn as acute (C1; V1) and chronic (C2; V2) ; all rats of acute and chronic groups were sacrificed under the anesthesia on the 45th day for acute group [C1 (n=5) and V1 (n=12)] and the 90th day for chronic group [C2 (n=5) and V2 (n=12)], respectively. Thus, biochemical and histopathological analysis were carried out. Results Thirty four rats completed the study, 24 were in varenicline group and 10 were in control group. In chronic exposure to varenicline, oxidant levels comprising of malondialdehyde (MDA), and myeloperoxidase (MPO) increased and superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) and glutathione peroxidase (GPx) levels, named as antioxidants, decreased significantly when compared to the control group. MDA and MPO levels were also significantly higher and SOD, CAT, GPx, GSH levels were also significantly lower in chronic varenicline group when compared to acute varenicline group. These findings were also supported by histopathological observations. Conclusion This is the first study, which evaluated pulmonary effects of varenicline experimentally on an animal model. It was observed that chronic varenicline treatments cause inflammation and lung cell injury.

Estudo da morfologia e status oxidativo em rim de ratos com nefropatia isquêmica após tratamento com células-tronco mesenquimais de tecido adiposo / Study of morphology and oxidative status in kidney of rats with ischemic nephropathy after treatment with adipose tissue mesenchymal stem cells

A nefropatia isquêmica é uma doença renal crônica provocada pela redução do fluxo sanguíneo renal que pode progredir para a doença renal terminal, cujo tratamentos disponíveis se baseiam em terapias substitutivas da função renal, como diálise ou transplante renal. No entanto, devido ao alto custo dos tratamentos e a carência de órgãos, se faz necessária a busca por novas terapias, como as células-tronco (CT). Apesar do potencial terapêutico das CT em doenças crônicas, não está claro se essas células mantêm seus efeitos benéficos em órgãos lesionados por tempo prolongado. O objetivo desse estudo foi avaliar os efeitos precoces e tardios do tratamento com células-tronco adiposas (CTA) sobre a morfologia e o status oxidativo em rins de ratos com nefropatia isquêmica. A isquemia renal foi induzida pelo modelo 2rins-1clip (2R1C) e, depois de um mês da clipagem da artéria renal, foram injetadas 106 células-tronco na região subscapsular do rim afetado. Após 15 e 30 dias da injeção das CTA, a morfologia renal foi verificada por meio da análise macroscópica, microscópica e ultraestrutural. Além disso, o status oxidativo foi avaliado no tecido renal através da mensuração da atividade das enzimas antioxidantes catalase e glutationa peroxidase; e de marcadores biológicos de dano oxidativo, como proteínas carboniladas, 3-Nitrotirosina e 4-Hidroxinonenal. Por imunoperoxidase foi possível localizar as células-tronco adiposas GFP+ foram rastreadas e encontradas tanto 15 dias, quanto 30 dias após a injeção na região subcapsular. A restauração da arquitetura renal foi evidenciada 15d após o uso das células, onde detectamos redução na deposição de fibras colágenas no parênquima renal, o que não foi observado 30d após o uso das células. Os resultados também foram confirmados através da análise da ultraestrutura renal que mostraram restauração da arquitetura renal no grupo de 15d, não evidenciada no grupo de 30d. Quanto a análise do status oxidativo, somente os animais com nefropatia isquêmica mais prolongada apresentaram estresse oxidativo com redução da atividade da enzima antioxidante catalase no tecido renal. Além disso, foi observado dano proteico e lipídico, sem melhora dessa condição nos animais 30d após o tratamento com as células-tronco. No modelo de nefropatia isquêmica avaliado, o tratamento com CTA mostrou benefícios na morfologia renal a curto prazo, mas não tardiamente, apesar da permanência dessas células no tecido. Acreditamos que o estresse oxidativo, evidenciado somente no tecido renal com isquemia mais prolongada, possa ter dificultado a ação das células-tronco, contribuindo para tais achados. Esses resultados abrem perspectivas para o aprofundamento do estudo quanto à caracterização dos mecanimos de ação das CTA nas respostas anti-fibrogênicas, assim como o estabelecimento do número, frequência, vias de administração e melhor momento para uso dessas células no tratamento de doenças renais crônicas.

Ischemic nephropathy is a chronic kidney disease caused by reduced kidney blood flow that can progress to end stage kidney disease, whose available treatments are based on kidney function replacement therapies, such as dialysis or kidney transplantation. However, due to the high cost of treatments and the lack of organs, it is necessary to search for new therapies, such as stem cells (SC). Despite the therapeutic potential of SC in chronic diseases, it is unclear whether these cells maintain their beneficial effects on injured organs for a long time. The aim of this study was to evaluate the early and late effects of adipose-derived stem cells (ADSC) treatment on the morphology and oxidative status in kidneys of rats with ischemic nephropathy. Renal ischemia was induced by the 2kidneys-1clip (2K1C) model and, after a month of clipping the renal artery, 106 stem cells were injected into the subscapsular region of the affected kidney. After 15 and 30 days of ADSC injection, renal morphology was verified by macroscopic, microscopic, and ultrastructural analysis. In addition, oxidative status was assessed in renal tissue by measuring the activity of the antioxidant enzymes catalase and glutathione peroxidase; and biological markers of oxidative damage, such as carbonylated proteins, 3-nitrotyrosine and 4-hydroxynonenal. By immunoperoxidase, it was possible to locate GFP + adipose-derived stem cells that were tracked and found both 15 days and 30 days after injection in the subcapsular region. The restoration of the renal architecture was evidenced 15d after the use of the cells, where we detected a reduction in the deposition of collagen fibers in the renal parenchyma, which was not observed 30d after the use of the cells. The results were also confirmed by analyzing the renal ultrastructure, which showed restoration of the renal architecture in the 15d group, not evidenced in the 30d group. Regarding the analysis of oxidative status, only animals with more prolonged ischemic nephropathy presented oxidative stress with reduced activity of the antioxidant enzyme catalase in renal tissue. In addition, protein and lipid damage was observed, with no improvement in this condition in the animals 30d after treatment with stem cells. In the evaluated ischemic nephropathy model, treatment with ADSC showed benefits in renal morphology in the short term, but not late, despite the permanence of these cells in the tissue. We believe that oxidative stress, evidenced only in renal tissue with more prolonged ischemia, may have hindered the action of stem cells, contributing to such findings. These results open perspectives for further study on the characterization of ADSC mechanisms of action in anti-fibrogenic responses, as well as the establishment of the number, frequency, routes of administration and the best time to use these cells in the treatment of chronic kidney diseases.

miR-9-1 gene methylation and DNMT3B (rs2424913) polymorphism may contribute to periodontitis

Abstract Genetic and epigenetic changes have been associated with periodontitis in various genes; however, little is known about genes involved in epigenetic mechanisms and in oxidative stress. Objective: This study aims to investigate the association of polymorphisms C677T in MTHFR (rs1801133) and −149C→T in DNMT3B (rs2424913), as well as the methylation profiles of MTHFR, miR-9-1, miR-9-3, SOD1, and CAT with periodontitis. The association between polymorphisms and DNA methylation profiles was also analyzed. Methodology: The population studied was composed of 100 nonsmokers of both sexes, divided into healthy and periodontitis groups. Genomic DNA was extracted from the epithelial buccal cells, which were collected through a mouthwash. Polymorphism analysis was performed through polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), while methylation-specific PCR (MSP) or combined bisulfite restriction analysis techniques were applied for methylation analysis. Results: For DNMT3B, the T allele and the TT genotype were detected more frequently in the periodontitis group, as well as the methylated profile on the miR-9-1 promoter region. There was also a tendency towards promoter region methylation on the CAT sequence of individuals with periodontal disease. Conclusion: The polymorphism −149C→T in DNMT3B (rs2424913) and the methylated profile of the miR-9-1 promoter region are associated with periodontitis.