Criopreservação de células-tronco mesenquimais do tecido adiposo: Uma alternativa para lipoenxertia seriada na reconstrução mamária / Cryopreservation of adipose tissue-derived mesenchymal stem cells: An alternative to serial fat grafting in breast reconstruction

Introdução: A lipoenxertia é um enxerto autólogo de células do tecido celular subcutâneo, que pode ser utilizada como técnica complementar na reconstrução mamária. Diante disso, a criopreservação de células-tronco mesenquimais provenientes de tecido adiposo (CTDAs) poderia ser uma maneira de realizar a coleta em um tempo cirúrgico e após realizar a lipoenxertia de forma fracionada. O dimetilsulfóxido (DMSO) é um criopreservante utilizado em pesquisas com células, porém é potencialmente tóxico, o que impossibilitaria a utilização de CTDAs criopreservadas na prática clínica. Novos criopreservantes celulares, sem toxicidade, vêm sendo descritos na literatura científica experimental, como as substâncias L-prolina e trealose. Com isso, esse trabalho teve como objetivo avaliar a viabilidade de CTDAs criopreservadas com a combinação de L-prolina e trealose, em um período de até 90 dias. Método: Estudo experimental, no qual foram obtidas amostras de lipoaspirado provenientes de 9 pacientes. A fração celular foi processada e congelada com L-prolina (1,5M) + trealose (0,2M), ou com DMSO + soro fetal bovino (SFB), como controle. Após 30 e 90 dias, as amostras foram descongeladas e a viabilidade celular foi avaliada pela técnica de MTT. Resultados: A análise das CTDAs, após 1 e 3 meses de congelamento, indicou que as amostras tratadas com L-prolina + trealose apresentaram viabilidade semelhante àquelas preservadas com DMSO e SFB (p=0,444). Conclusão: A associação de L-prolina e trealose manteve CTDA viáveis por 30 e 90 dias de congelamento, podendo ser uma alternativa como criopreservante celular sem toxicidade e viabilizando o uso de lipoenxertia seriada.

Introduction: Fat grafting is an autologous graft of cells from subcutaneous tissue, which can be used as a complementary technique in breast reconstruction. Given this, the cryopreservation of adipose tissue-derived mesenchymal stem cells (ADMSCs) could be a way to collect them in one surgical procedure and after performing fractional fat grafting. Dimethyl sulfoxide (DMSO) is a cryopreservative used in cell research, but it is potentially toxic, which would make it impossible to use cryopreserved ADMSCs in clinical practice. New cellular cryopreservatives, without toxicity, have been described in the experimental scientific literature, such as the substances L-proline and trehalose. Therefore, this work aimed to evaluate the viability of ADMSCs cryopreserved with the combination of L-proline and trehalose over up to 90 days. Method: Experimental study in which lipoaspirate samples were obtained from 9 patients. The cellular fraction was processed and frozen with L-proline (1.5M) + trehalose (0.2M) or with DMSO + fetal bovine serum (FBS) as control. After 30 and 90 days, the samples were thawed, and cell viability was assessed using the MTT technique. Results: The analysis of ADMSCs, after 1 and 3 months of freezing, indicated that samples treated with L-proline + trehalose showed similar viability to those preserved with DMSO and SFB (p=0.444). Conclusion: The association of L-proline and trehalose kept ADMSC viable for 30 and 90 days of freezing, and could be an alternative as a cellular cryopreservative without toxicity and enabling the use of serial fat grafting.

Activation of M1 muscarinic acetylcholine receptors by proline-rich oligopeptide 7a (

Background: The bioactive peptides derived from snake venoms of the Viperidae family species have been promising as therapeutic candidates for neuroprotection due to their ability to prevent neuronal cell loss, injury, and death. Therefore, this study aimed to evaluate the cytoprotective effects of a synthetic proline-rich oligopeptide 7a (PRO-7a; <EDGPIPP) from Bothrops jararaca snake, on oxidative stress-induced toxicity in neuronal PC12 cells and astrocyte-like C6 cells. Methods: Both cells were pre-treated for four hours with different concentrations of PRO-7a, submitted to H2O2-induced damage for 20 h, and then the oxidative stress markers were analyzed. Also, two independent neuroprotective mechanisms were investigated: a) L-arginine metabolite generation via argininosuccinate synthetase (AsS) activity regulation to produce agmatine or polyamines with neuroprotective properties; b) M1 mAChR receptor subtype activation pathway to reduce oxidative stress and neuron injury. Results: PRO-7a was not cytoprotective in C6 cells, but potentiated the H2O2-induced damage to cell integrity at a concentration lower than 0.38 μM. However, PRO-7a at 1.56 µM, on the other hand, modified H2O2-induced toxicity in PC12 cells by restoring cell integrity, mitochondrial metabolism, ROS generation, and arginase indirect activity. The α-Methyl-DL-aspartic acid (MDLA) and L-NΩ-Nitroarginine methyl ester (L-Name), specific inhibitors of AsS and nitric oxide synthase (NOS), which catalyzes the synthesis of polyamines and NO from L-arginine, did not suppress PRO-7a-mediated cytoprotection against oxidative stress. It suggested that its mechanism is independent of the production of L-arginine metabolites with neuroprotective properties by increased AsS activity. On the other hand, the neuroprotective effect of PRO-7a was blocked in the presence of dicyclomine hydrochloride (DCH), an M1 mAChR antagonist. Conclusions: For the first time, this work provides evidence that PRO-7a-induced neuroprotection seems to be mediated through M1 mAChR activation in PC12 cells, which reduces oxidative stress independently of AsS activity and L-arginine bioavailability.(AU)

Factor inducible por hipoxia en cáncer

El oxígeno y dióxido de carbono son vitales en la respiración, sus variaciones fuera del rango fisiológico son una amenaza para la supervivencia de las células. La hipoxia es una condición común en la mayoría de los tumores malignos, la cual promueve angiogénesis y vascularización disfuncional, mayor proliferación celular y la adquisición de un fenotipo de transición epitelial a mesenquimatoso, que contribuye con la metástasis; asimismo, altera el metabolismo de las células cancerosas y genera resistencia a la terapia, ya que induce a la inactividad celular. Por tanto, la hipoxia es un factor negativo, asociado a resultados adversos en la mayoría de los tratamientos de los distintos tipos de cáncer. El factor inducible por hipoxia (HIF) es el factor de transcripción relacionado con la hipoxia en cáncer, que produce la activación de más de una centena de genes reguladores de la actividad celular, que generan funciones cruciales para el desarrollo del cáncer. El objetivo principal de la presente revisión es puntualizar la importancia de la hipoxia en la génesis del cáncer, conocer las principales moléculas que interactúan en la expresión del HIF, explicar los mecanismos moleculares de las vías involucradas en la inducción del HIF, las consecuencias celulares por su alteración y las potenciales terapias dirigidas contra este factor. Se consultaron PubMed, Scopus y SciELO, del año 1990 hasta el año 2022, y se buscaron las referencias bibliográficas en relación con las palabras clave asociadas al factor inducible por hipoxia y cáncer. En conclusión, la sobreexpresión de HIF-1α en biopsias tumorales se asocia con una mayor mortalidad de pacientes en cánceres humanos. Los posibles genes diana regulados por HIF-1α que pueden desempeñar un papel en la progresión tumoral están empezando a descubrirse. A pesar de que se han estudiado cientos de compuestos en relación con el HIF en cáncer, en la actualidad existen pocos inhibidores del HIF aprobados en el mercado mundial; asimismo, muchos estudios clínicos, en sus distintas fases en desarrollo, no muestran resultados alentadores. Probablemente, en el futuro, cuando se tenga una mejor comprensión de la estructura, funcionamiento molecular y biológico de este factor, se desarrollarán fármacos más específicos para la inhibición del HIF.

Oxygen and carbon dioxide are essential for breathing; variations in these gases outside of the normal range are a threat to cell survival. Hypoxia is a common condition that occurs in most malignant tumors, increases angiogenesis and defective vascularization, promotes cell proliferation and acquires an epithelial-mesenchymal transition phenotype, which causes metastasis. It also affects cancer cell metabolism and makes patients resistant to treatment by causing cell quiescence. As a result, hypoxia is a detrimental component that is linked to unfavorable outcomes in most cancer treatments. Through the activation of more than a hundred genes that control cell activity, which produce key functions for cancer development, the transcription factor known as hypoxia-inducible factor (HIF) is linked to hypoxia in cancer. This review's main goals are to highlight the role of hypoxia in the development of cancer, identify the key molecules that interact to promote HIF expression, explain the molecular mechanisms of the pathways that lead to HIF induction, describe the cellular effects of HIF alteration, and discuss potential HIF-targeted therapies. Articles from 1990 to 2022 were reviewed in PubMed, Scopus and SciELO databases. Keywords related to cancer and HIF were searched in bibliographical references. In conclusion, HIF-1α overexpression in tumor biopsies is associated with increased patient mortality in human cancers. Potential HIF-1α-regulated target genes that may play a role in tumor progression are starting to be identified. Although hundreds of chemicals have been studied in relation to HIF in cancer, there are currently few approved HIF inhibitors available on the global market; moreover, many clinical trials, in their various stages of development, do not show encouraging results. It is likely that in the future, when there is a better understanding of the structure, molecular and biological functioning of this factor, more specific drugs for HIF inhibition will be developed.

Osmoregulatory Appraisal of Some Osmoprotectants on Hydrolytic Activities of Some Enzymes on Seeds of Two Water Stressed Cultivars of Sorghum bicolor (Moench)

Enzymes play significant roles in metabolic processes of seeds. Therefore, this study evaluated osmoregulatory potential of some osmoprotectants on activities of some hydrolytic enzymes in the seeds of two cultivars (SOSAT.C-88 and CV. LCIC 9702) of sorghum bicolor. Matured seeds of the two cultivars were harvested and prepared for alpha, beta, total amylase and proteinase activities assay. The osmoprotectants produced significant variations on the enzymes at 10 and 14 days (DA) of 8 weeks after treatments (WAT). Seeds of well-watered SOSAT.C-88 produced higher alpha (2.10 IU/ml), beta (1.70 IU/ml) and total amylase activities (3.30 IU/ml) at 14 days (DA). Higher alpha (2.01 IU/ml and total amylase activities (2.61 IU/ml) were recorded in the seeds of CV. LCIC 9702 well-watered at 14 days DA 8WAT. Furthermore, total amylase activities (3.87 IU/ml) were recorded in the seeds produced by CV. LCIC 9702 well-watered at 14 days DA. Significant increase was noticed in beta (1.14 IU/ml) and alpha amylase (1.58 IU/ml) in the seeds of CV. LCIC 9702 treated with mycorrhiza. CV. LCIC 9702 well watered produced highest proteinase activities (1.57 U/ml) while least of the parameters were recorded in SOSAT.C-88 and CV. LCIC 9702 droughted. In conclusion, the osmoprotectants had regulatory effects on the activities of hydrolytic enzymes therefore the use of the osmoprotectants in farming should be encouraged.

Meta-analysis of Ac-SDKP inhibition of Pulmonary fibrosis in animal models / 中华劳动卫生职业病杂志

Objective: To systematically study the anti-fibrotic effect of N-acetyl-seryl-as partyl-lysyl-proline (Ac-SDKP) on pulmonary fibrosis. Methods: In May 2021, a computer search was performed on CNKI, Wanfang Knowledge Service Platform, VIP.com, China Biomedical Literature Database, Pubmed, OVID and other databases. The retrieval time was from January 2008 to May 2021. Randomized controlled experiments on the inhibition of pulmonary fibrosis by Ac-SDKP were screened. The control group was the pulmonary fibrosis model group and the experimental group was the Ac-SDKP treatment group. The quality of the literature was assessed using the syrcle risk of bias assessment tool, and data were extracted. Data analysis was Performed using revman 5.4 software. Results: 18 papers were included, with a total of 428 animal models. The results of meta analysis showed that the contents of α-smooth muscle actin (α-SMA), type I collagen, type Ⅲ collagen, transforming growth factor-β (TGF-β) and Nodule area in the exPerimental group were lower than those in the control grouP. [SMD=-2.44, 95%CI (-3.71--1.17), P=0.000][SMD=-5.36, 95%CI (-7.13--3.59), P=0.000] [SMD=-3.07, 95%CI (-4.13--2.02), P<0.000][SMD=-2.88, 95%CI (-3.63--2.14), P=0.000] [SMD=-1.80, 95%CI (-2.42--1.18), P=0.000], the content of hydroxy proline in the experimental group was higher than that in the control group [SMD=7.62, 95%CI (4.90-10.33), P=0.000], all indexes included in the literature were statistically significant. Conclusion: Ac-SDKP has obvious inhibitory effect on the process of pulmonary fibrosis, and may become a new clinical drug for the treatment of pulmonary fibrosis.

Physiochemical response of papaya genotypes exposed to low temperature regimes

Susceptibility to low temperature stress is the major threat to papaya cultivation. Here, we studied a low temperature stress tolerance in papaya plant. We have investigated the effect of different low temperature regimes, 28°/18°C (day/night) to 16°/06°C (day/night) with a gradual decrease of 2°C on every two days and one set with direct exposure to the low temperature of 18°/08°C (day/night), called the acclimatized plant, in five diverse papaya genotypes (Pusa Nanha, Red Lady P-7-2, P-7-9, and P-7-14) and cold tolerant wild relative of cultivated papaya genotype (Vasconcellea cundinamarcensis V.M. Badillo) under controlled regulated conditions. It was observed that there were significant variations in the physiological and biochemical parameters like photosynthetic gas exchange parameters, chlorophyll content, fluorescence parameters, relative water content (RWC), membrane stability index (MSI), total sugars content, total soluble proteins content, lipid peroxidation, and proline accumulation in leaf tissues. Maximum stomatal conductance, chlorophyll fluorescence, RWC, MSI, total sugars, total soluble proteins, proline and lowest MDA contents were observed in Vasconcellea cundinamarcensis followed by inbred P-7-9 as compared to other genotypes under low temperature stress. Among all the papaya genotypes, P-7-9 showed more adaptability to low temperature stress and it further give new insights for developing low temperature tolerant papaya genotypes, especially under changing climate situations.

Production of proline and protease with different organic wastes in bacteria (Production proline and protease with organic wastes) / Produção de prolina e protease com diferentes resíduos orgânicos em bactérias (produção de prolina e protease com resíduos orgânicos)

In this study, we investigated the proline and protease production of different bacteria in several organic waste materials. Our aim was to produce proline and protease economically in waste that is abundantly available while reducing its environmental impact. 5 ml of different organic waste materials (OWW: Olive waste water; N.B: Nutrient Broth; EW: Eggshell; PBS: PBS buffer; PLW: Peach leaf wastes; TCW: Turkish coffee wastes; TWW: Tea waste water; WCW: Waste cheese whey; WFO: Waste frying oil) were placed in 10 ml grow tubes, inoculated and incubated for 24 h. Phosphate-buffered saline and 10% solutions of different organic wastes were added. These cultures were subsequently incubated at 37°C for 24 h. Cells were harvested at 24 h for L-proline assay. 1 ml of culture was transferred by pipette into an Eppendorf tube and centrifuged at 14,000 rpm for 20 min at room temperature. Cellular debris was removed by centrifuge and the supernatant was used for proline activity assays. Protease activity was determined using a modified method with casein as the substrate. We found that proline and protease can easily be produced economically using Turkish coffee wastes (TCW), Waste cheese whey (WCW) and Olive waste water (OWW) organic waste. We believe that this study will result in similar research leading to the economical use of these waste materials thus reducing their impact on the environment.

Neste estudo, investigamos a produção de prolina e protease de diferentes bactérias em diversos resíduos orgânicos. Nosso objetivo era produzir prolina e protease economicamente em resíduos que estão disponíveis em abundância, reduzindo seu impacto ambiental. Cinco ml de diferentes materiais de resíduos orgânicos (OWW: resíduos de azeitona; NB: caldo nutriente; EW: casca de ovo; PBS: tampão PBS; PLW: resíduos de folhas de pêssego; TCW: resíduos de café turco; TWW: resíduos de chá; WCW: resíduos de queijo soro de leite; WFO: óleo de fritura residual) foram colocados em tubos de cultivo de 10 ml, inoculados e incubados por 24 horas. Adicionaram-se solução salina tamponada com fosfato e soluções a 10% de diferentes resíduos orgânicos. Essas culturas foram subsequentemente incubadas a 37° C durante 24 h. As células foram colhidas às 24 h para o ensaio de L-prolina. Um ml de cultura foi transferido por pipeta para um tubo Eppendorf e centrifugado a 14.000 rpm, por 20 min, em temperatura ambiente. Os detritos celulares foram removidos por centrifugação e o sobrenadante foi usado para ensaios de atividade de prolina. A atividade da protease foi determinada usando um método modificado com caseína como substrato. Descobrimos que a prolina e a protease podem ser facilmente produzidas economicamente, usando resíduos de café turco (TCW), resíduos de soro de queijo (WCW) e resíduos orgânicos de água de oliva (OWW). Acreditamos que este estudo resultará em pesquisas semelhantes, levando ao uso econômico desses materiais residuais, reduzindo, assim, seu impacto no meio ambiente.

Expression of proline rich protein 11 in breast cancer and its relationship with clinical biological behavior, prognosis and survival / 中华内分泌外科杂志

Objective:To study the expression of Proline rich protein11 (PRR11) in breast cancer and its relationship with clinical biological behavior, prognosis and survival.Methods:A prospective analysis method was used to select 80 patients with breast cancer from Jan. 2018 to Jan. 2019. Immunohistochemical S-P method was used to detect the expression of PRR11 in cancer tissues. Patients with positive expression of PRR11 were set as the study group ( n=47) and the patients with negative expression of PRR11 were set as the control group ( n=33) . All patients were followed up for 3 years to analyze and compare the survival rates of patients with positive and negative expression of PRR11. The relationship between PRR11 expression and clinical biological behavior, prognosis and survival was analyzed by Cox risk ratio review model. Results:80 patients were followed up for 3 years. It was found that the prognosis of patients with negative PRR11 expression was significantly better than that of patients with positive PRR11 expression ( χ2=5.75, P<0.001) . Chi square test was used to analyze the correlation between the expression of PRR11 and tumor size, TNM stage, lymph node metastasis, distant metastasis, histological grade, Ki67 expression and hormone receptor status ( P<0.05) . The expression of PRR11 in breast cancer tissues with larger tumors, distant metastasis and later staging was relatively high ( P<0.05) . Univariate Cox regression analysis showed that histological grade, TNM stage and PRR11 were independent risk factors affecting the prognosis of breast cancer patients ( P<0.001) . The AUC of prognosis prediction in patients with breast cancer was 0.812, and the 95% CI was 0.635-0.796. When PRR11 expression was positive, the sensitivity was 81.47%, and the specificity was 85.57%. Conclusions:The expression of PRR11 is relatively high in the late stage breast cancer tissue. The expression of PRR11 is closely related to the clinical biological behavior of breast cancer size, TNM stage and lymph node metastasis. The survival rate of patients with high PRR11 expression is low, and the positive expression of PRR11 is an independent risk factor affecting the prognosis of breast cancer patients. PRR11 detection has preferable clinical application value in predicting the prognosis of breast cancer.

Plasma metabolomics study in patients with occupational silicosis / 中国职业医学

@#Abstract: Objective To analyze the plasma metabolomics of patients with occupational silicosis, and screen the differential Methods metabolic pathways in different stages of silicosis. Thirty patients with occupational silicosis were selected as silicosis group by judgment sampling method, including 10 patients in each subgroups with silicosis at stages Ⅰ, Ⅱ, and Ⅲ. Another 10 healthy individuals without occupational dust exposure history were selected as the control group. Plasma of each patientwascollected.Themetabolitesfromtheirplasmasampleswerecollectedandidentifiedbyuntargetedmetabolomicsusing----ultraperformanceliquidchromatographyquadrupoletimeofflightmassspectrometry.Principalcomponentanalysisandpartial least square discriminant analysis were used to compare the differentially expressed metabolites. The metabolic pathways Results enrichment analysis was performed using MetaboAnanlyst 5.0 and other metabolic analysis software. There were significantlydifferentmetabolicprofilesamongtheplasmaofthesilicosissubgroupsandthecontrolgroup.Inthepositivemode,--149differentmetaboliteswerescreened,amongwhich99metaboliteswereupregulatedand50metabolitesweredown regulated. Sphingolipids metabolism, arginine and ornithine metabolism, and fatty acid degradation are the main metabolic pathwaysinsilicosissubgroupswithstagesⅠandⅡcomparedwiththecontrolgroup.Themetabolicpathwaysofsilicosisstage Ⅲ are arginine and proline metabolism, glycerol metabolism, glyoxylic acid and dicarboxylic acid metabolism, and glycine, Conclusion serine and threonine metabolism. The plasma metabolic profile of patients with silicosis in different stages changed significantly. Sphingolipid metabolism is the main metabolic pathway in the early and middle stages of silicosis. In - silicosisterminalstage,thechangeofarginineandproline basedmetabolicpathwayresultinginfibrosisaggravation.

Advances on microbial synthesis of L-proline and trans-4-hydroxy-L-proline / 生物工程学报

L-proline (L-Pro) is the only imino acid among the 20 amino acids that constitute biological proteins, and its main hydroxylated product is trans-4-hydroxy-L-proline (T-4-Hyp). Both of them have unique biological activities and play important roles in biomedicine, food and beauty industry. With the in-depth exploration of the functions of L-Pro and T-4-Hyp, the demand for them is gradually increasing. Traditional methods of biological extraction and chemical synthesis are unable to meet the demand of "green, environmental protection and high efficiency". In recent years, synthetic biology has developed rapidly. Through the intensive analysis of the synthetic pathways of L-Pro and T-4-Hyp, microbial cell factories were constructed for large-scale production, which opened a new chapter for the green and efficient production of L-Pro and T-4-Hyp. This paper reviews the application and production methods of L-Pro and T-4-Hyp, the metabolic pathways for microbial synthesis of L-Pro and T-4-Hyp, and the engineering strategies and advances on microbial production of L-Pro and T-4-Hyp, aiming to provide a theoretical basis for the "green bio-manufacturing" of L-Pro and T-4-Hyp and promote their industrial production.

Antioxidant status and their enhancements strategies for water stress tolerance in chickpea / Status antioxidante e suas estratégias de aprimoramento para tolerância ao estresse hídrico no grão-de-bico

Water stress executes severe influences on the plant growth and development through modifying physio-chemical properties. Therefore, a field experiment was designed to evaluate the antioxidant status and their enhancements strategies for water stress tolerance in chickpea on loam and clay loam soils under agro-ecological conditions of Arid Zone Research Institute, Bahawalpur (29.3871 °N, 71.653 °E) and Cholistan farm near Derawer (28.19°N, 71.80°E) of Southern Punjab, Pakistan during winter 2014-15. Experimental treatments comprised of two chickpea cultivars i.e. Bhakhar 2011 (drought tolerant) and DUSHT (drought sensitive), two water stress levels i.e. water stress at flowering stage and water stress at flowering + pod formation + grain filling stage including well watered (control) and two exogenous application of osmoprotectants i.e. glycine betaine (GB) 20 ppm and proline 10 uM including distilled water (control). Results indicated that water stress at various growth stages adversely affects the growth, yield and quality attributes of both chickpea cultivars. Exogenous application of GB and proline improved the growth, yield and quality parameters of both chickpea cultivars even under water stress conditions. However, superior results were obtained with exogenously applied GB on Bhakhar 2011 under well-watered conditions. Similarly, foliar spray of GB on chickpea cultivar Bhakhar 2011 under stress at flowering + pod formation + grain filling stage produced maximum superoxide dismutase, peroxidase and catalase contents. These results suggested that application of GB mitigates the adverse effects of water stress and enhanced tolerance in chickpea mainly due to higher antioxidant enzymes activity, demonstrating the protective measures of plant cells in stress condition. Hence, antioxidants status might be a suitable method for illustrating water stress tolerance in chickpea.

O estresse hídrico exerce fortes influências no crescimento e no desenvolvimento das plantas, modificando as propriedades físico-químicas. Portanto, a presente atividade de pesquisa foi projetada para avaliar o status antioxidante e suas estratégias de aprimoramento para tolerância ao estresse hídrico no grão-de-bico em condições agroecológicas, no Instituto de Pesquisa da Zona Árida, Bahawalpur (29.3871 ° N, 71.653 ° E) e fazenda do Cholistan, perto de Derawer (28.19 ° N, 71,80 ° E), no sul de Punjab, Paquistão, durante Rabi 2014-15. Tratamentos experimentais compostos de dois genótipos de grão-de-bico, como Bhakhar 2011 (tolerante à seca) e DUSHT (sensível à seca), dois níveis de estresse hídrico, ou seja, estresse hídrico no estágio de floração, estresse hídrico na fase de floração e estresse hídrico na fase de floração + formação de vagem + estágio de enchimento de grãos, incluindo água bem controlada (controle) e duas aplicações exógenas de osmoprotetores, isto é, glicina betaína 20 ppm e prolina 10 uM, incluindo água destilada (controle). Os resultados indicaram que o estresse hídrico em vários estágios de crescimento afeta negativamente os atributos de crescimento, rendimento e qualidade de ambas as cultivares de grão-de-bico. A aplicação exógena de glicina betaína e prolina melhorou os parâmetros de crescimento, rendimento e qualidade de ambos os genótipos de grão- de-bico, mesmo sob condições de estresse hídrico. No entanto, resultados superiores foram obtidos com glicina betaína aplicada exogenamente em Bhakhar 2011, em condições bem regadas. Além disso, o spray foliar de glicina betaína na cultivar de grão-de-bico Bhakhar 2011, sob estresse na fase de floração + formação de vagem + enchimento de grãos, produziu o máximo de superóxido dismutase, peroxidase e catalase. Esses resultados sugeriram que a aplicação de glicina betaína atenua os efeitos adversos do estresse hídrico e aumenta a tolerância no grão-de- bico, principalmente pela maior atividade de enzimas antioxidantes, demonstrando medidas protetoras das células vegetais em condições de estresse. Portanto, o status de antioxidantes pode ser um método adequado para ilustrar a tolerância ao estresse hídrico no grão-de-bico.

Antioxidant status and their enhancements strategies for water stress tolerance in chickpea / Status antioxidante e suas estratégias de aprimoramento para tolerância ao estresse hídrico no grão-de-bico

Water stress executes severe influences on the plant growth and development through modifying physio-chemical properties. Therefore, a field experiment was designed to evaluate the antioxidant status and their enhancements strategies for water stress tolerance in chickpea on loam and clay loam soils under agro-ecological conditions of Arid Zone Research Institute, Bahawalpur (29.3871 °N, 71.653 °E) and Cholistan farm near Derawer (28.19°N, 71.80°E) of Southern Punjab, Pakistan during winter 2014-15. Experimental treatments comprised of two chickpea cultivars i.e. Bhakhar 2011 (drought tolerant) and DUSHT (drought sensitive), two water stress levels i.e. water stress at flowering stage and water stress at flowering + pod formation + grain filling stage including well watered (control) and two exogenous application of osmoprotectants i.e. glycine betaine (GB) 20 ppm and proline 10 uM including distilled water (control). Results indicated that water stress at various growth stages adversely affects the growth, yield and quality attributes of both chickpea cultivars. Exogenous application of GB and proline improved the growth, yield and quality parameters of both chickpea cultivars even under water stress conditions. However, superior results were obtained with exogenously applied GB on Bhakhar 2011 under well-watered conditions. Similarly, foliar spray of GB on chickpea cultivar Bhakhar 2011 under stress at flowering + pod formation + grain filling stage produced maximum superoxide dismutase, peroxidase and catalase contents. These results suggested that application of GB mitigates the adverse effects of water stress and enhanced tolerance in chickpea mainly due to higher antioxidant enzymes activity, demonstrating the protective measures of plant cells in stress condition. Hence, antioxidants status might be a suitable method for illustrating water stress tolerance in chickpea.

O estresse hídrico exerce fortes influências no crescimento e no desenvolvimento das plantas, modificando as propriedades físico-químicas. Portanto, a presente atividade de pesquisa foi projetada para avaliar o status antioxidante e suas estratégias de aprimoramento para tolerância ao estresse hídrico no grão-de-bico em condiçõesa groecológicas, no Instituto de Pesquisa da Zona Árida, Bahawalpur (29.3871 ° N, 71.653 ° E) e fazenda do Cholistan, perto de Derawer (28.19 ° N, 71,80 ° E), no sul de Punjab, Paquistão, durante Rabi 2014-15. Tratamentos experimentais compostos de dois genótipos de grão-de-bico, como Bhakhar 2011 (tolerante à seca) e DUSHT (sensível à seca), dois níveis de estresse hídrico, ou seja, estresse hídrico no estágio de floração, estresse hídrico na fase de floração e estresse hídrico na fase de floração + formação de vagem + estágio de enchimento de grãos, incluindo água bem controlada (controle) e duas aplicações exógenas de osmoprotetores, isto é, glicina betaína 20 ppm e prolina 10 uM, incluindo água destilada (controle). Os resultados indicaram que o estresse hídrico em vários estágios de crescimento afeta negativamente os atributos de crescimento, rendimento e qualidade de ambas as cultivares de grão-de-bico. A aplicação exógena de glicina betaína e prolina melhorou os parâmetros de crescimento, rendimento e qualidade de ambos os genótipos de grão- de-bico, mesmo sob condições de estresse hídrico. No entanto, resultados superiores foram obtidos com glicina betaína aplicada exogenamente em Bhakhar 2011, em condições bem regadas. Além disso, o spray foliar de glicina betaína na cultivar de grão-de-bico Bhakhar 2011, sob estresse na fase de floração + formação de vagem + enchimento de grãos, produziu o máximo de superóxido dismutase, peroxidase e catalase. Esses resultados sugeriram que a aplicação de glicina betaína atenua os efeitos adversos do estresse hídrico e aumenta a [...].

Production of proline and protease with different organic wastes in bacteria (Production proline and protease with organic wastes) / Produção de prolina e protease com diferentes resíduos orgânicos em bactérias (produção de prolina e protease com resíduos orgânicos)

In this study, we investigated the proline and protease production of different bacteria in several organic waste materials. Our aim was to produce proline and protease economically in waste that is abundantly available while reducing its environmental impact. 5 ml of different organic waste materials (OWW: Olive waste water; N.B: Nutrient Broth; EW: Eggshell; PBS: PBS buffer; PLW: Peach leaf wastes; TCW: Turkish coffee wastes; TWW: Tea waste water; WCW: Waste cheese whey; WFO: Waste frying oil) were placed in 10 ml grow tubes, inoculated and incubated for 24 h. Phosphate-buffered saline and 10% solutions of different organic wastes were added. These cultures were subsequently incubated at 37°C for 24 h. Cells were harvested at 24 h for L-proline assay. 1 ml of culture was transferred by pipette into an Eppendorf tube and centrifuged at 14,000 rpm for 20 min at room temperature. Cellular debris was removed by centrifuge and the supernatant was used for proline activity assays. Protease activity was determined using a modified method with casein as the substrate. We found that proline and protease can easily be produced economically using Turkish coffee wastes (TCW), Waste cheese whey (WCW) and Olive waste water (OWW) organic waste. We believe that this study will result in similar research leading to the economical use of these waste materials thus reducing their impact on the environment.

Neste estudo, investigamos a produção de prolina e protease de diferentes bactérias em diversos resíduos orgânicos. Nosso objetivo era produzir prolina e protease economicamente em resíduos que estão disponíveis em abundância, reduzindo seu impacto ambiental. Cinco ml de diferentes materiais de resíduos orgânicos (OWW: resíduos de azeitona; NB: caldo nutriente; EW: casca de ovo; PBS: tampão PBS; PLW: resíduos de folhas de pêssego; TCW: resíduos de café turco; TWW: resíduos de chá; WCW: resíduos de queijo soro de leite; WFO: óleo de fritura residual) foram colocados em tubos de cultivo de 10 ml, inoculados e incubados por 24 horas. Adicionaram-se solução salina tamponada com fosfato e soluções a 10% de diferentes resíduos orgânicos. Essas culturas foram subsequentemente incubadas a 37° C durante 24 h. As células foram colhidas às 24 h para o ensaio de L-prolina. Um ml de cultura foi transferido por pipeta para um tubo Eppendorf e centrifugado a 14.000 rpm, por 20 min, em temperatura ambiente. Os detritos celulares foram removidos por centrifugação e o sobrenadante foi usado para ensaios de atividade de prolina. A atividade da protease foi determinada usando um método modificado com caseína como substrato. Descobrimos que a prolina e a protease podem ser facilmente produzidas economicamente, usando resíduos de café turco (TCW), resíduos de soro de queijo (WCW) e resíduos orgânicos de água de oliva (OWW). Acreditamos que este estudo resultará em pesquisas semelhantes, levando ao uso econômico desses materiais residuais, reduzindo, assim, seu impacto no meio ambiente.

Antioxidant status and their enhancements strategies for water stress tolerance in chickpea

Abstract Water stress executes severe influences on the plant growth and development through modifying physio-chemical properties. Therefore, a field experiment was designed to evaluate the antioxidant status and their enhancements strategies for water stress tolerance in chickpea on loam and clay loam soils under agro-ecological conditions of Arid Zone Research Institute, Bahawalpur (29.3871 °N, 71.653 °E) and Cholistan farm near Derawer (28.19°N, 71.80°E) of Southern Punjab, Pakistan during winter 2014-15. Experimental treatments comprised of two chickpea cultivars i.e. Bhakhar 2011 (drought tolerant) and DUSHT (drought sensitive), two water stress levels i.e. water stress at flowering stage and water stress at flowering + pod formation + grain filling stage including well watered (control) and two exogenous application of osmoprotectants i.e. glycine betaine (GB) 20 ppm and proline 10 uM including distilled water (control). Results indicated that water stress at various growth stages adversely affects the growth, yield and quality attributes of both chickpea cultivars. Exogenous application of GB and proline improved the growth, yield and quality parameters of both chickpea cultivars even under water stress conditions. However, superior results were obtained with exogenously applied GB on Bhakhar 2011 under well-watered conditions. Similarly, foliar spray of GB on chickpea cultivar Bhakhar 2011 under stress at flowering + pod formation + grain filling stage produced maximum superoxide dismutase, peroxidase and catalase contents. These results suggested that application of GB mitigates the adverse effects of water stress and enhanced tolerance in chickpea mainly due to higher antioxidant enzymes activity, demonstrating the protective measures of plant cells in stress condition. Hence, antioxidants status might be a suitable method for illustrating water stress tolerance in chickpea.

Resumo O estresse hídrico exerce fortes influências no crescimento e no desenvolvimento das plantas, modificando as propriedades físico-químicas. Portanto, a presente atividade de pesquisa foi projetada para avaliar o status antioxidante e suas estratégias de aprimoramento para tolerância ao estresse hídrico no grão-de-bico em condições agroecológicas, no Instituto de Pesquisa da Zona Árida, Bahawalpur (29.3871 ° N, 71.653 ° E) e fazenda do Cholistan, perto de Derawer (28.19 ° N, 71,80 ° E), no sul de Punjab, Paquistão, durante Rabi 2014-15. Tratamentos experimentais compostos de dois genótipos de grão-de-bico, como Bhakhar 2011 (tolerante à seca) e DUSHT (sensível à seca), dois níveis de estresse hídrico, ou seja, estresse hídrico no estágio de floração, estresse hídrico na fase de floração e estresse hídrico na fase de floração + formação de vagem + estágio de enchimento de grãos, incluindo água bem controlada (controle) e duas aplicações exógenas de osmoprotetores, isto é, glicina betaína 20 ppm e prolina 10 uM, incluindo água destilada (controle). Os resultados indicaram que o estresse hídrico em vários estágios de crescimento afeta negativamente os atributos de crescimento, rendimento e qualidade de ambas as cultivares de grão-de-bico. A aplicação exógena de glicina betaína e prolina melhorou os parâmetros de crescimento, rendimento e qualidade de ambos os genótipos de grão- de-bico, mesmo sob condições de estresse hídrico. No entanto, resultados superiores foram obtidos com glicina betaína aplicada exogenamente em Bhakhar 2011, em condições bem regadas. Além disso, o spray foliar de glicina betaína na cultivar de grão-de-bico Bhakhar 2011, sob estresse na fase de floração + formação de vagem + enchimento de grãos, produziu o máximo de superóxido dismutase, peroxidase e catalase. Esses resultados sugeriram que a aplicação de glicina betaína atenua os efeitos adversos do estresse hídrico e aumenta a tolerância no grão-de- bico, principalmente pela maior atividade de enzimas antioxidantes, demonstrando medidas protetoras das células vegetais em condições de estresse. Portanto, o status de antioxidantes pode ser um método adequado para ilustrar a tolerância ao estresse hídrico no grão-de-bico.

Production of proline and protease with different organic wastes in bacteria (Production proline and protease with organic wastes)

Abstract In this study, we investigated the proline and protease production of different bacteria in several organic waste materials. Our aim was to produce proline and protease economically in waste that is abundantly available while reducing its environmental impact. 5 ml of different organic waste materials (OWW: Olive waste water; N.B: Nutrient Broth; EW: Eggshell; PBS: PBS buffer; PLW: Peach leaf wastes; TCW: Turkish coffee wastes; TWW: Tea waste water; WCW: Waste cheese whey; WFO: Waste frying oil) were placed in 10 ml grow tubes, inoculated and incubated for 24 h. Phosphate-buffered saline and 10% solutions of different organic wastes were added. These cultures were subsequently incubated at 37°C for 24 h. Cells were harvested at 24 h for L-proline assay. 1 ml of culture was transferred by pipette into an Eppendorf tube and centrifuged at 14,000 rpm for 20 min at room temperature. Cellular debris was removed by centrifuge and the supernatant was used for proline activity assays. Protease activity was determined using a modified method with casein as the substrate. We found that proline and protease can easily be produced economically using Turkish coffee wastes (TCW), Waste cheese whey (WCW) and Olive waste water (OWW) organic waste. We believe that this study will result in similar research leading to the economical use of these waste materials thus reducing their impact on the environment.

Resumo Neste estudo, investigamos a produção de prolina e protease de diferentes bactérias em diversos resíduos orgânicos. Nosso objetivo era produzir prolina e protease economicamente em resíduos que estão disponíveis em abundância, reduzindo seu impacto ambiental. Cinco ml de diferentes materiais de resíduos orgânicos (OWW: resíduos de azeitona; NB: caldo nutriente; EW: casca de ovo; PBS: tampão PBS; PLW: resíduos de folhas de pêssego; TCW: resíduos de café turco; TWW: resíduos de chá; WCW: resíduos de queijo soro de leite; WFO: óleo de fritura residual) foram colocados em tubos de cultivo de 10 ml, inoculados e incubados por 24 horas. Adicionaram-se solução salina tamponada com fosfato e soluções a 10% de diferentes resíduos orgânicos. Essas culturas foram subsequentemente incubadas a 37° C durante 24 h. As células foram colhidas às 24 h para o ensaio de L-prolina. Um ml de cultura foi transferido por pipeta para um tubo Eppendorf e centrifugado a 14.000 rpm, por 20 min, em temperatura ambiente. Os detritos celulares foram removidos por centrifugação e o sobrenadante foi usado para ensaios de atividade de prolina. A atividade da protease foi determinada usando um método modificado com caseína como substrato. Descobrimos que a prolina e a protease podem ser facilmente produzidas economicamente, usando resíduos de café turco (TCW), resíduos de soro de queijo (WCW) e resíduos orgânicos de água de oliva (OWW). Acreditamos que este estudo resultará em pesquisas semelhantes, levando ao uso econômico desses materiais residuais, reduzindo, assim, seu impacto no meio ambiente.

Regulatory effect of Ac-SDKP on phosphorylated heat shock protein 27/SNAI1 pathway in silicotic rats / 中华劳动卫生职业病杂志

Objective: To study the effect of anti-fibrotic tetrapeptide N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) on phosphorylated heat shock protein 27 (P-HSP27) and zinc finger family transcriptional repressor 1 (SNAI1) expression to explore the anti-silicosis fibrosis effect of Ac-SDKP. Methods: In December 2014, the rat silicosis animal model was prepared by one-time bronchial infusion of silicon dioxide (SiO(2)) dust. 80 SPF healthy adult Wistar rats were selected, and the rats were divided into 8 groups according to the random number table method, 10 in each group. Model control group for 4 weeks (feeding for 4 weeks) , model control group for 8 weeks (feeding for 8 weeks) : bronchial perfusion with normal saline 1.0 ml per animal. Silicosis model group for 4 weeks (feeding for 4 weeks) and silicosis model group for 8 weeks (feeding for 8 weeks) : bronchial perfusion of 50 mg/ml SiO(2) suspension 1.0 ml per animal. Ac-SDKP administration group for 4 weeks (feeding for 4 weeks) , Ac-SDKP administration group for 8 weeks (feeding for 8 weeks) : Ac-SDKP 800 μg·kg(-1)·d(-1) was administered by intraperitoneal pump. Ac-SDKP preventive treatment group: 48 h after Ac-SDKP 800 μg·kg(-1)·d(-1) administration, bronchial perfusion of SiO(2) suspension 1.0 ml per animal, raised for 8 weeks. Ac-SDKP anti-fibrosis treatment group: after bronchial perfusion of 1.0 ml of SiO(2) suspension for 4 weeks, Ac-SDKP 800 μg·kg(-1)·d(-1) was administered for 4 weeks. Western blotting was used to detect the expression of P-HSP27, SNAI1, α-smooth muscle actin (α-SMA) , and collage typeⅠ and Ⅲ in each group. The expression of P-HSP27 and SNAI1 was detected by immunohistochemistry, and the co-localized expression of P-HSP27 and α-SMA was detected by laser confocal microscopy. Results: Compared with the model control group, the expressions of P-HSP27, SNAI1, α-SMA, and collage typeⅠ and Ⅲ in the silicosis fibrosis area of the rats in the silicosis model group were enhanced, and the differences were statistically significant (P<0.05) . After Ac-SDKP intervention, compared with silicosis model group for 8 weeks, the expressions of P-HSP27, SNAI1 α-SMA, and collage typeⅠ and Ⅲ in the Ac-SDKP preventive and anti-fibrosis treatment groups were significantly decreased, and the differences were statistically significant (P<0.05) . However, the expressions of P-HSP27 SNAI1, and collage typeⅠ and Ⅲ between the Ac-SDKP administration group and the model control group did not change significantly, and the differences were not statistically significant (P>0.05) . Laser confocal results showed that the positive cells expressing P-HSP27 and α-SMA in the lung tissue of the silicosis model group were more than those in the model control group. Compared with the silicosis model group, the Ac-SDKP prevention and anti-fibrosis treatment groups expressing the positive cells of P-HSP27 and α-SMA decreased. Compared with the model control group for 8 weeks, there were some double-positive cells expressing P-HSP27 and α-SMA in the nodules of the silicosis model group for 8 weeks. Conclusion: Ac-SDKP may play an anti-silicic fibrosis effect by regulating the P-HSP27/SNAI1 pathway.

Construction and application of a synthetic promoter library for Corynebacterium glutamicum / 生物工程学报

Promoter is an important genetic tool for fine-tuning of gene expression and has been widely used for metabolic engineering. Corynebacterium glutamicum is an important chassis for industrial biotechnology. However, promoter libraries that are applicable to C. glutamicum have been rarely reported, except for a few developed based on synthetic sequences containing random mutations. In this study, we constructed a promoter library based on the native promoter of odhA gene by mutating the -10 region and the bystanders. Using a red fluorescent protein (RFP) as the reporter, 57 promoter mutants were screened by fluorescence imaging technology in a high-throughput manner. These mutants spanned a strength range between 2.4-fold and 19.6-fold improvements of the wild-type promoter. The strongest mutant exhibited a 2.3-fold higher strength than the widely used strong inducible promoter Ptrc. Sequencing of all 57 mutants revealed that 55 mutants share a 1-4 bases shift (4 bases shift for 68% mutants) of the conserved -10 motif "TANNNT" to the 3' end of the promoter, compared to the wild-type promoter. Conserved T or G bases at different positions were observed for strong, moderate, and weak promoter mutants. Finally, five promoter mutants with different strength were employed to fine-tune the expression of γ-glutamyl kinase (ProB) for L-proline biosynthesis. Increased promoter strength led to enhanced L-proline production and the highest L-proline titer of 6.4 g/L was obtained when a promoter mutant with a 9.8-fold higher strength compared to the wild-type promoter was used for ProB expression. The use of stronger promoter variants did not further improve L-proline production. In conclusion, a promoter library was constructed based on a native C. glutamicum promoter PodhA. The new promoter library should be useful for systems metabolic engineering of C. glutamicum. The strategy of mutating native promoter may also guide the construction of promoter libraries for other microorganisms.

Expression of PRR11 in Bladder Cancer Tissue and Its Effect on Proliferation and Apoptosis of Bladder Cancer Cell Line T24 / 肿瘤防治研究

Objective To investigate the expression of PRR11 in bladder cancer tissues and its effect on proliferation and apoptosis of bladder cancer cell line T24. Methods The expression of PRR11 was detected using immunohistochemistry method in 57 specimens of bladder urothelial carcinoma and adjacent tissues. The correlations of PRR11 expression with the clinicopathological characteristics of patients with bladder urothelial carcinoma were analyzed. The mRNA and protein expression levels of PRR11 in human immortalized bladder epithelial cell lines SV-HUC-1 and human bladder cancer cell lines HTB-9, T24, J82 and UM-UC-3 were measured by qRT-PCR and Western blot. The gene expression of PRR11 in T24 cells was silenced by lentivirus shRNA. The mRNA expression level of PRR11 was detected by qRT-PCR. CCK-8 was used to detect cell proliferative activity. Cell clonality was detected by plate cloning assays. The rate of apoptosis was evaluated using flow cytometry. The protein expression levels of PRR11, Caspase-3, Bcl-2 and Bax were assessed by Western blot. Results PRR11 was highly expressed in bladder urothelial carcinoma, and its expression level was correlated with the pathological grade and T stage of the tumor. The mRNA and protein expression levels of PRR11 in HTB-9, T24, J82 and UM-UC-3 cells were higher than those in SV-HUC-1 cells (P < 0.05), especially in T24 cells. PRR11 gene silence reduced the expression levels of PRR11 mRNA and protein, as well as the cell proliferation activity and cell clonality, elevated the apoptosis rate, up-regulated the protein expression levels of Cleaved caspase-3 and Bax, and down-regulated the protein expression level of Bcl-2. Conclusion The expression of PRR11 is upregulated in bladder urothelial carcinoma tissues and bladder cancer cell lines. Interfering with PRR11 expression can inhibit the proliferation and promote the apoptosis of bladder cancer T24 cells.

Arabidopsis CK2 family gene CKB3 involved in abscisic acid signaling / Gene da família Arabidopsis CK2 CKB3 envolvido na sinalização de ácido abscísico

CKB3 is a regulatory (beta) subunit of CK2. In this study Arabidopsis thaliana homozygous T-DNA mutant ckb3 was studied to understand the role of CKB3 in abscisic acid (ABA) signaling. The results shown: CKB3 was expressed in all organs and the highest expression in the seeds, followed by the root. During seed germination and root growth the ckb3 mutant showed reduced sensitivity to ABA. The ckb3 mutant had more stomatal opening and increased proline accumulation and leaf water loss. The expression levels of number of genes in the ABA regulatory network had changed. This study demonstrates that CKB3 is an ABA signaling-related gene and may play a positive role in ABA signaling.

CKB3 é uma subunidade reguladora (beta) de CK2. Neste estudo, o mutante homozigoto ckb3 de Arabidopsis thaliana foi estudado para entender o papel da CKB3 na sinalização de ácido abscísico (ABA). Os resultados apresentados: CKB3 foi expresso em todos os órgãos e a maior expressão nas sementes, seguida pela raiz. Durante a germinação das sementes e o crescimento radicular, o mutante ckb3 mostrou sensibilidade reduzida ao ABA. O mutante ckb3 teve mais abertura estomática e aumento do acúmulo de prolina e perda de água nas folhas. Os níveis de expressão do número de genes na rede reguladora da ABA haviam mudado. Este estudo demonstra que CKB3 é um gene relacionado à sinalização ABA e pode desempenhar um papel positivo na sinalização ABA.

Infuence of kaolin application on most important fruit and leaf characteristics of two apple cultivars under sustained defcit irrigation

BACKGROUND: Apple is one of the oldest and most valuable fruits. Water restriction is one of the major problems in the production of this fruit in some planting areas. METHODS: Effects of kaolin spray treatments were studied on two early apple cultivars of Golab and Shafi-Abadi under sustained deficit irrigation (SDI) in Alborz province, Iran during 2017 and 2018. Irrigation treatments were 100%, 85%, and 70% ETc and kaolin application were concentrations of 0, 3 and 6% in 2017 and 0, 1.5 and 3% in 2018. RESULTS: Results showed that 85% ETc treatment compared to other irrigation treatments improved apple tree crown volume in 2017. Deficit irrigation treatments significantly reduced fruit weight in both years. Application with 6% kaolin resulted in 33.3% increase in apple fruit weight compared to non-kaolin treatment at 100% ETc irrigation in the first year. Severe deficit irrigation (70% ETc) significantly reduced apple fruit length in both years, but 6% kaolin increased fruit length in both apple cultivars in 2017. Severe deficit irrigation treatment increased the firmness of apple fruit compared to control and mild deficit irrigation (85% ETc) in the first year of experiment. There was no significant difference between irrigation treatments for apple fruit firmness in the second year of experiment. Kaolin treatments of 1.5% and 3% at full irrigation increased the soluble solids content of apple fruit by 36.6% and 44.1% in 2018, respectively. Deficit irrigation treatments significantly increased leaf proline content compared to control in both years. In the first year, kaolin treatments increased leaf proline but in the second year, leaf proline was not significant. Deficit irrigation treatment of 70% ETc and 6% kaolin had the highest amount of glycine betaine content, malondialdehyde and hydrogen peroxide in apple leaf in the first year of experiment. CONCLUSIONS: Severe deficit irrigation stress (70% ETc) increased the activity of nonenzymatic defense systems of apple trees. Kaolin as a drought stress reducing agent can be recommended in apple orchards of Golab and Shafi-Abadi cultivars as an effective and inexpensive method to improve tolerance to drought stress conditions.