Delta-aminolevulinic Acid Dehydratase Research Articles

Engineered production of 5-aminolevulinic acid in recombinant Escherichia coli BL21.

5-aminolevulinic acid (ALA) is a non-protein amino acid that has been widely used in the fields of medicine and agriculture. This study aims to engineer the C5 pathway of the ALA biosynthesis in Escherichia coli BL21 to enhance ALA production. The ALA synthase genes gltX, hemA, and hemL were overexpressed in E. coli BL21 to lead to the increase in the production of ALA. The sRNA RyhB was also overexpressed to downregulate the expression of ALA dehydratase to reduce the downstream bioconversion of ALA to porphobilinogen. Next, the gene arcA was knocked out by CRISPR-Cas9 technology to open the TCA cycle to promote the respiratory metabolism of the strain to reduce the feedback inhibition of heme to ALA. The fermentation conditions of the engineered strain were optimized by response surface experiments. The time-course analysis of the ALA production was carried out in a 1 L shake flask. Through these efforts, the production of ALA in engineered strain reached 2953 mg/L in a 1 L shake flask. This study contributes to the industrial production of ALA by the engineered E. coli in the future.

Low temperature and high humidity affect dynamics of chlorophyll biosynthesis and secondary metabolites in Cucumber.

During the cold season, low temperature (LT) and high relative humidity (HRH) are significant environmental factors in greenhouses and plastic tunnels, often hindering plant growth and development. The chlorophyll (Chl) biosynthesis inhibitory mechanisms under LT and HRH stress are still widely unclear. To understand how cucumbers seedlings respond to LT and HRH stress, we investigated the impact of these stressors on Chl biosynthesis. Our results revealed that individual LT, HRH and combined LT + HRH stress conditions affected chlorophyll a, b, total chlorophyll and carotenoid content, reducing the levels of these pigments. The levels of Chlorophyll precursors were also markedly reduced under LT and HRH stresses, with the greatest reduction observed in cucumber seedlings exposed to LT + HRH conditions (9/5℃, 95%HRH). The activities of glutamate-1-semialdehyde transaminase (GSA-AT), ALA dehydratase (ALAD), Mg-chelatase, and protochlorophyllide oxidoreductase (POR) were increased under individual LT, HRH, conditions but decreased by combination of LT + HRH stress condition. In addition, Chl biosynthesis related genes (except PBG) were upregulated by the HRH stress but were significantly downregulated under the LT + HRH stress condition in cucumber seedlings. Furthermore, the content of phenols, flavonoids and phenolic acids (cinnamic acid and caffeic acid) were significantly surged under LT + HRH treatment over the control. Histochemical observation showed higher O2- and H2O2 content in cucumber leaves during the LT and HRH stress. The results indicate that LT + HRH stress significantly impairs chlorophyll biosynthesis in cucumber seedlings by drastically reducing pigment accumulation, altering enzyme activity and gene expression. Additionally, LT + HRH stress induces oxidative damage, which further exacerbates the decline in chlorophyll content and affects overall cucumber metabolism.

Delta-Aminolevulinic acid dehydratase enzyme activity and susceptibility to lead toxicity in Uganda’s urban children

Background With rapid industrialization, urbanization, and population explosion in sub-Saharan Africa including Uganda, the population has experienced increased exposure to environmental lead subsequently causing elevated blood lead levels. Mean blood levels of 332µg/dL,120µg/dʟ, 25µg/dL,11µg/dL, and 10µg/dL in children under 18 years of age in Nigeria, DR Congo, South Africa, Sudan, and Uganda respectively. Susceptibility to lead toxicity correlates with one’s nutrition status, age, and genetics. This study expounded susceptibility to lead toxicity by relating blood lead levels, delta-aminolevulinic acid dehydratase (ALAD) enzyme activity, and genetic variations of proteins that code for ALAD in urban children of Uganda aged between 6 and 60 months. Methods A total of 198 blood samples were analyzed for blood lead levels (BLL), on an atomic absorption spectrophotometer whereas hemoglobin (Hb) levels, and ALAD enzyme activity, were analyzed on a spectrophotometer before DNA extraction, polymerase chain reaction, and restriction fragment length digestion for ALAD polymorphism. Results Geometric means of BLL (10.55µg/dL, SD = 7.4), Hb (7.85g/dL, SD = 1.3) and ALAD enzyme activity (37.15 units/L BLL, S.D = 9.7), corresponded to samples that coded for ALAD1 allele (99.05%) compared to the 0.05% that coded for ALAD2 with BLL (14.5µg/ dL, SD = 4.7), Hb (6.1 g/ dL), ALAD enzyme activity (33.8 units/L, SD=1.45). There was a significant relationship with a negative linear correlation between BLL, Hb (status, and ALAD enzyme activity in the three isozymes (ALAD1-1, ALAD1-2, and ALAD2-2) in the strength of ALAD1-1 (r = 0.42, p-value = 0.02) ˂ ALAD1-2 (r = 0.62, effective size = 0.43, p-value = ˂ 0.001) ˂ ALAD2-2 (r = 0.67, effective size = 0.86, p-value = ˂ 0.001). Conclusions Most of the study participants coded for the ALAD1 allele hence hoarded blood lead, which could result in delayed exposure and adverse effects later in their lives.

Changes in Metabolism and Content of Chlorophyll in Common Duckweed (Lemna minor L.) Caused by Environmental Contamination with Fluorides.

The impact of fluorine on plants remains poorly understood. We examined duckweed growth in extracts of soil contaminated with fluorine leached from chicken manure. Additionally, fluorine levels were analyzed in fresh manure, outdoor-stored manure, and soil samples at varying distances from the manure pile. Fresh manure contained 37-48 mg F- × kg-1, while soil extracts contained 2.1 to 4.9 mg F- × kg-1. We evaluated the physiological effects of fluorine on duckweed cultured on soil extracts or in 50% Murashige-Skoog (MS) medium supplemented with fluorine concentrations matching those in soil samples (2.1 to 4.9 mg F- × L-1), as well as at 0, 4, and 210 mg × L-1. Duckweed exposed to fluorine displayed similar toxicity symptoms whether in soil extracts or supplemented medium. Fluoride at concentrations of 2.1 to 4.9 mg F- × L-1 reduced the intact chlorophyll content, binding the porphyrin ring at position 32 without affecting Mg2+. This reaction resulted in chlorophyll a absorption peak shifted towards shorter wavelengths and formation of a new band of the F--chlorophyll a complex at λ = 421 nm. Moreover, plants exposed to low concentrations of fluorine exhibited increased activities of aminolevulinic acid dehydratase and chlorophyllase, whereas the activities of both enzymes sharply declined when the fluoride concentration exceeded 4.9 mg × L-1. Consequently, fluorine damages chlorophyll a, disrupts the activity of chlorophyll-metabolizing enzymes, and diminishes the plant growth rate, even when the effects of these disruptions are too subtle to be discerned by the naked human eye.

DNA damage and ALAD polymorphism in high blood lead (Pb) levels of pregnant women attending a tertiary care teaching hospital

Pregnant women are particularly vulnerable to lead toxicity due to increased absorption and decreased elimination of lead from their bodies. The δ-aminolevulinic acid dehydratase (ALAD) gene plays a crucial role in lead metabolism, and its polymorphisms have been implicated in modifying the susceptibility to lead toxicity. A cross-sectional study was conducted involving 90 pregnant women and blood samples were collected to measure blood lead levels (BLL) and assessed DNA damage using the comet assay. ALAD polymorphisms were genotyped using PCR-RFLP analysis with MspI restriction enzyme. Statistical analysis, including chi-square tests, logistic regression, and correlation analysis, was performed to determine associations between ALAD polymorphisms, BLL, and DNA damage. From 90 pregnant women the participants, 16 had high BLL (≥5μg/dL), while the remaining 74 had normal levels (<5μg/dL). The ALAD 1-2 genotype was found to be significantly associated with high BLL (p<0.001). Pregnant women with the ALAD 1-2 genotype exhibited higher levels of DNA damage compared to those with other genotypes (p<0.001). Furthermore, a positive correlation was observed between the transfer of lead concentration from mother to infant and DNA damage severity (r=0.511, p<0.001). The combination of comet assay and polymorphism analysis offers a comprehensive approach to understanding the impact of lead exposure during pregnancy. These findings underscore the urgent need for effective regulatory measures to reduce lead exposure in the environment and mitigate its adverse effects of lead on maternal and child health.

Abstract B046: Irradiation of patient-derived high-grade gliomas reduces 5-ALA-induced fluorescence utility during re-resection

Abstract High grade gliomas HGGs are among the most aggressive primary brain tumors, yielding a median survival time of less than 2 years and a 5-year survival of 2.5%. There has been little in the way of treatments and novel approaches are needed to combat HGG’s poor prognosis. Recent studies have established that HGG cells exhibit metabolic reprogramming to adapt to diverse metabolic gradients within heterogenous tumor microenvironments. Using an unbiased metabolomics approach, we investigated metabolic changes both pre- and post-ionizing radiation across several patient-derived cell lines. Acute dosages of ionizing radiation resulted in significant changes in the synthesis of aminolevulinic acid (ALA), which is known to be converted into the fluorescent metabolite protoporphyrin IX (PpIX) in the porphyrin synthesis pathway. PpIX has been utilized by surgeons in fluorescence-guided tumor resection for clearer visualization of tumor boundaries in recent years. Fractionation of radiation therapy also resulted in dose-dependent changes in the pathway concerning heme synthesis within these cells, especially presenting decreased levels of ALA dehydratase (ALAD) and increased levels of ferrochelatase. As ALAD promotes PpIX accumulation and ferrochelatase catalyzes its conversion to heme, this indicates a decreased favorability for the accumulation of PpIX in the reprogrammed metabolism of HGG cells following radiation. Consequently, we propose that PpIX fluorescence is not a reliable visual aide in surgical re-resection of HGGs post-radiation therapy. Citation Format: Paras S. Minhas, Aarooran Durairaj, Melanie McReynolds, Katrin I. Andreasson, YooJin Jung, Joshua Rabinowitz. Irradiation of patient-derived high-grade gliomas reduces 5-ALA-induced fluorescence utility during re-resection [abstract]. In: Proceedings of the AACR Special Conference on Brain Cancer; 2023 Oct 19-22; Minneapolis, Minnesota. Philadelphia (PA): AACR; Cancer Res 2024;84(5 Suppl_1):Abstract nr B046.

The role of laboratory diagnostics in the assessment of occupational lead exposure

Introduction: Industrialization and urbanization led to a significant increase in the environment. Lead inhibits the activity of numerous enzymes, triggers oxidative stress, and causes protein biosynthesis dysregulation. Inhalation of lead particles is the most common route of intoxication associated with occupational exposure. This study aims to evaluate laboratory methods and biomarkers in the assessment of lead exposure. Methods: For non-experimental qualitative research, available scientific articles in English published in the relevant databases (MEDLINE and ScienceDirect) were used. The database search was performed using the keywords “Laboratory diagnostics”, “occupational exposure”, and “lead”. Results: Atomic absorption spectrometry (AAS) is the gold standard in laboratory monitoring of occupational lead exposure. Inductively coupled plasma with mass spectrometry is a commonly used method described as more sensitive than AAS due to its low detection limit. Lead concentrations can be determined in various samples, but blood and urine are the most commonly used in laboratory practice. The most important exposure biomarker is the enzyme δ-aminolevulinic acid dehydratase (ALAD) in the blood, which is characterized by progressive inactivation by lead and a negative correlation with its concentration. The concentration of urinary delta-aminolevulinic acid (δ-ALA-U) reflects the state of impaired enzyme function in heme biosynthesis. In addition, determining blood zinc protoporphyrin and urinary coproporphyrin levels significantly aids in assessing occupational lead exposure disorders. Conclusion: The availability of the laboratory methods used and the biomarker specificity and sensitivity play an important role in the adequacy of lead exposure monitoring. Accurate determination of ALAD and δ-ALA-U concentrations, along with other biomarkers, is critical for assessing individuals exposed to lead.

Impact of hemB mutations on 5-aminolevulinic acid production in Escherichia coli

Background Microbial production of 5-aminolevulinic acid (ALA) attracts attention due to a wide range of biotechnological and medical applications of ALA, including cancer treatment and diagnosis. Various genetic engineering approaches have been employed to improve ALA production in bacterial hosts such as Escherichia coli possessing the C5 pathway. Glutamyl-tRNA reductase (GluTR) encoded by hemA, glutamate-1-semialdehyde aminotransferase (GSA-AT) encoded by hemL, and ALA dehydratase (ALAD) encoded by hemB play important roles in ALA metabolism including the C5 pathway. Attenuation of the intercellular ALAD activity, which condensates 2 molecules of ALA to synthesize porphobilinogen (PBG), has been employed by various measures. However, a mutation approach by substituting catalytically important residues in ALAD encoded by hemB has never been attempted. The aim of this study is to assess the impact of hemB mutations on the ALA production in E. coli. Methods In this study, the authors mutated the amino acid residues potentially related to the enzymatic activity of E. coli ALAD by referring to a mutation experiment of human ALAD. The authors created five types of mutated hemB genes, introduced these genes to the hemB-deleted mutant strain of E. coli, and assessed the impact of the ALAD mutations on ALA production. In addition, hemA, hemL, and rhtA encoding an ALA exporter were introduced to the E. coli possessing a mutated hemB. Results The authors revealed that the mutations of ALAD employed in this study did not significantly enhance ALA production. Overexpression of hemA, hemL, and rhtA substantially increased ALA production in any E. coli strain possessing mutated hemB, while a difference in ALA production of the strain could be rather attributed to its growth behaviour than ALAD inactivation. Conclusions This study provides an important piece of information to design the bioprocess of ALA production using E. coli engineered through the C5 pathway.

Potassium and jasmonic acid —Induced nitrogen and sulfur metabolisms improve resilience against arsenate toxicity in tomato seedlings

Despite the well-known individual roles of potassium (K+) and jasmonic acid (JA) in plant responses to various stresses, their combined effects on improving tolerance to metalloid arsenate (AsV) toxicity and their influence on physiological and biochemical mechanisms remain largely unexplored. Therefore, the present study was conducted to explore the concomitant role of K+ and JA in nitrogen (N) and sulfur (S) metabolisms and regulation of antioxidant system in tomato seedlings under AsV toxicity conditions. The obtained data reveal that K+ and/or JA boosted the tolerance of tomato (Solanum lycopersicum L.) seedlings to AsV toxicity. However, the effect of a combined dose of K+ and JA was found to be more efficient as compared to the alone application. Seedlings subjected to AsV exhibited suppressed morphological attributes (shoot and root length, shoot and root fresh weight, and shoot and root dry weight), and physio-biochemical characteristics. Also, AsV-treated tomato seedlings showed enhanced reactive oxygen species (ROS) and increased the activity of ROS-producing enzymes (O2•— -generating NADPH oxidase activity and H2O2-generating glycolate oxidase] and also a chlorophyll (Chl) degrading enzyme [chlorophyllase (Chlase) activity]. However, the exogenous supply of K+ and JA synergistically inhibited Chl degradation and overproduction of ROS by suppressing their responsible enzymes activity. Moreover, it induced gas exchange parameters and activity of enzymes responsible for the photosynthesis process (carbonic anhydrase and Rubisco), Chl biosynthesis (δ-aminolevulinic acid dehydratase), and suppressed Chlase activity, and overproduction of ROS and their producing enzymes activity. Most interestingly, the combined application of K+ and JA substantially increased N and S content and their assimilation by upregulating the activity of enzymes involved in these key pathways (nitrate reductase, nitrite reductase, glutamine synthetase, glutamate synthase, ATP sulfurylase, adenosine 5-phosphosulfate reductase, O-acetylserine (thiol) lyase and sulfide reductase). The obtained results strongly reveal that the combined application of K+ and JA improved tolerance of tomato seedlings to AsV toxicity by enhancing N and S assimilation pathways and antioxidant system.

Strigolactone and nitric oxide collaborate synergistically to boost tomato seedling resilience to arsenic toxicity via modulating physiology and antioxidant system

Arsenic (As) poses a significant environmental threat as a metalloid toxin, adversely affecting the health of both plants and animals. Strigolactones (SL) and nitric oxide (NO) are known to play crucial roles in plant physiology. Therefore, the present experiment was designed to investigate the potential cumulative role of SL (GR24–0.20 μM) and NO (100 μM) in mitigating the adverse effect of AsV (53 μM) by modulating physiological mechanisms in two genotypes of tomato (Riogrand and Super Strain 8). A sample randomized design with four replicates was used to arrange the experimental pots in the growth chamber. 45-d old both tomato cultivars under AsV toxicity exhibited reduced morphological attributes (root and shoot length, root and shoot fresh weight, and root and shoot dry weight) and physiological and biochemical characteristics [chlorophyll (Chl) a and b content, activity of δ-aminolevulinic acid dehydratase activity (an enzyme responsible for Chl biosynthesis), and carbonic anhydrase activity (an enzyme responsible for photosynthesis), and enhanced Chl degradation, overproduction of reactive oxygen species (ROS) and lipid peroxidation due to enhanced malondialdehyde (MDA) content. However, the combined application of SL and NO was more effective in enhancing the tolerance of both varieties to AsV toxicity compared to individual application. The combined application of SL and NO improved growth parameters, biosynthesis of Chls, NO and proline. However, the combined application significantly suppressed cellular damage by inhibiting MDA and overproduction of ROS in leaves and roots, as confirmed by the fluorescent microscopy study and markedly upregulated the antioxidant enzymes (catalase, peroxidase, superoxide dismutase, ascorbate dismutase and glutathione reductase) activity. This study provides clear evidence that the combined application of SL and NO supplementation significantly improves the resilience of tomato seedlings against AsV toxicity. The synergistic effect of SL and NO was confirmed by the application of cPTIO (an NO scavenger) with SL and NO. However, further molecular studies could be imperative to conclusively validate the simultaneous role of SL and NO in enhancing plant tolerance to abiotic stress.

Effect of Pb-exposure and B vitamin deficiencies on δ-aminolevulinic acid dehydratase activity among workers from Pb recycling plants.

Previous studies reported that Pb exposure causes a negative association with delta-aminolevulinic acid dehydratase activity (δ-ALAD), but the impact of Pb exposure (dose and time), B vitamin deficiencies, and lifestyle factors needs to be explored. In this study, the impact of Pb exposure, B vitamin deficiencies, and lifestyle factors on δ-ALAD activity among workers exposed to Pb from the Pb-recycling process was evaluated. Blood lead levels (BLLs), B vitamins (B6, B9, and B12), hematological factors (Hb% and HCT), lifestyle factors, and δ-ALAD activity was assessed in 170 male Pb-exposed workers engaged in the Pb recycling process. BLLs are estimated using the ICP-OES method. B vitamins in serum samples from workers were determined using the ELISA method. The δ-ALAD activity in whole blood samples was determined using the spectrophotometer method. The lifestyle factors were collected using a standard questionnaire. The δ-ALAD activity was significantly decreased in workers with the habits of alcohol use, tobacco consumption, hematocrit < 41%, mild and moderate categories of anemia, vitamin B6 and B12 deficiency, and BLL categories of 10-30, 30-50, and > 50µg/dL. Multiple regression analysis revealed that the independent variables of alcohol consumption (β = -0.170; P = 0.025), BLLs (β = -0.589; P = 0.001) and Hb% (β = 0.183; P = 0.001) significantly influenced the δ-ALAD activity with 44.2% (R2 = 0.442). Among the workers exposed to Pb from the Pb recycling plant, δ-ALAD activity was considerably reduced by Pb exposure, B vitamin deficiency, hematological parameters, and lifestyle factors.

Evaluation of the antimalarial properties of Solanum incanum L. leaf extract fractions and its ability to downregulate delta aminolevulinate dehydratase to prevent the establishment of malaria infection

Ethnopharmacological relevanceSolanum incanum L. is commonly used in traditional herbal medicine (THM) in Kenya for treating various ailments. Recent developments in disease treatment have introduced the concept of host-directed therapy (HDT). This approach involves targeting factors within the host cell that can impede the growth or replication of a pathogen. One such host factor is delta aminolevulinate dehydratase (δ-ALAD), the second enzyme in the heme biosynthesis pathway utilized by Plasmodium for growth. Studies using mice models have shown an increase in δ-ALAD expression during Plasmodium berghei infection. Another plant in the Solanum genus, S. guaranticum, has been found to inhibit δ-ALAD in red blood cells in vitro and in the brain in vivo. Is it possible that the bioactive compounds in S. incanum extracts could also be effective in HDT for malaria treatment? Aim of studyTo better assess the effectiveness of S. incanum leaf extracts as a curative and prophylaxis in malaria parasite infection, and to test the plant’s ability to decrease δ-ALAD expression. Materials and methodsThe leaves of S. incanum were collected, dried, and pulverized before being subjected to a successive extraction protocol to obtain crude, hexane, ethyl acetate, and aqueous extract fractions. Phytochemical analysis was conducted on all extract fractions, followed by GC-MS analysis of the fraction with the most potent antimalarial activity. An acute toxicity study was also performed on the extracted fractions. The potency of the extract fractions as curative and prophylactic antimalarial was then evaluated in THM using Plasmodium berghei-infected mice at a dose of 100 mg/kg. The extract fraction with the highest activity was further evaluated at varying doses and its effect on δ-ALAD was measured using RT-qPCR. The percentage of parasitemia and chemosuppression, and mean survival time were used as indices of activity. ResultsPhytochemical analysis revealed that the ethyl acetate and aqueous extract fractions contained high terpenoids, flavonoids, and phenols levels. However, alkaloids were only present in moderate quantities in the aqueous extract, and quinones were found in high levels only in the crude extract. Additionally, all extract fractions contained saponins in high levels but lacked tannins. While the plant extracts were found to be non-toxic, they did not exhibit curative antimalarial activity. However, all extract fractions showed prophylactic antimalarial activity, with the ethyl acetate extract having the highest percentage of chemosuppression even at doses of 250 and 1000 mg/kg. In the negative control, the expression of δ-ALAD was 5.4-fold, but this was significantly reduced to 2.3-fold when mice were treated with 250 mg/kg of the ethyl acetate fraction. GC-MS analysis of the ethyl acetate fraction revealed high percentages of 2-methyloctacosane, tetracosane, and decane. ConclusionThe fractions extracted from S. incanum leaves have been found to possess only antimalarial prophylactic properties, with the ethyl acetate extract fraction showing the most effective results. The activity of this fraction may be attributed to its ability to decrease the expression of δ-ALAD, as it contains an alkane compound implicated with enzyme-inhibitory activity.

Genetic susceptibility to low-level lead exposure in men: Insights from ALAD polymorphisms

The genetic susceptibility to low-level lead (Pb) exposure in general populations has been poorly investigated and is limited to the single nucleotide polymorphism (SNP) rs1800435 in the delta-aminolevulinic acid dehydratase gene (ALAD). This study explored associations between ten selected ALAD SNPs with Pb concentrations in blood (BPb) and urine (UPb) among 281 men aged 18–49 years from Slovenia, including 20 individuals residing in a Pb-contaminated area. The geometric mean (range) of BPb and UPb were 19.6 (3.86–84.7) μg/L and 0.69 (0.09–3.82) μg/L SG, respectively. The possible genetic influence was assessed by examining SNP haplotypes, individual SNPs, and the combination of two SNPs using multiple linear regression analyses. While no significant associations were found for haplotypes, the presence of variant alleles of rs1800435 and rs1805312 resulted in an 11% and 13% decrease in BPb, respectively, while the presence of variant allele of rs1139488 (homozygous only) exhibited significant 20% increase in BPb, respectively. Additionally, variant allele of rs1800435 resulted in lower UPb. Individual SNPs in the model explained only around 1 additional percentage point of BPb variability. In contrast, combination analyses identified six combinations of two SNPs, which significantly explained 3–22 additional percentage points of BPb variability, with the highest explanatory power observed for the rs1800435-rs1139488 and rs1139488-rs1805313 combinations. Moreover, excluding participants from the Pb-contaminated area indicated that exposure level influenced SNPs-Pb associations.Our results confirm the importance of the ALAD gene in Pb kinetics even at low exposure levels. Additionally, we demonstrated that identifying individuals with specific combinations of ALAD SNPs explained a larger part of Pb variability, suggesting that these combinations, pending confirmation in other populations and further evaluation through mechanistic studies, may serve as superior susceptibility biomarker in Pb exposure compared to individual SNPs.

S-nitrosoglutathione-facilitated activation of ATP synthase involves hydrogen sulfide during the response of plants to cadmium toxicity

The contamination of soils with cadmium (Cd) threatens agricultural productivity worldwide. The ability of plants to withstand challenging circumstances is primarily contingent upon their inherent capacity for tolerance and defense. The purpose of the current work is to gain a better understanding of how the mitochondrial ATP synthase enzyme, the antioxidant defense system, redox homeostasis, and chlorophyll metabolism in tomato seedlings are regulated by nitric oxide and hydrogen sulfide under Cd stress. The results demonstrate that exposure of tomato seedlings to Cd stress resulted in a substantial decrease in the activity of ATP synthase. The plants exposed to Cd stress, exhibited an increase in chlorophyllase activity and a substantial decrease in the activity of δ-aminolevulinic acid dehydratase, which resulted in a substantial loss of chlorophyll content. Elevation in the antioxidant defense system under Cd stress could not bring a significant positive change in the plants’ defense system that resulted in lipid peroxidation and leakage of electrolytes. Incubation of Cd-stressed seedlings with nitric oxide donor S-nitrosoglutathione (GSNO) proved remarkable in improving the performance of the plants. The results exhibit that GSNO application improved the activity of ATP synthase, superoxide dismutase, peroxidase, and catalase and the performance of the ascorbate-glutathione system, which efficiently scavenged excess reactive oxygen species. The positive effect of GSNO application was also shown by decreased lipid peroxidation and electrolyte leakage. However, application of DL-propargylglycine (a biosynthesis inhibitor of hydrogen sulfide) to the stressed-seedlings reversed the effect of GSNO, which resulted in the weak performance of the defense system and the commencement of oxidative stress. The results of the study substantiate that nitric oxide, in association with endogenous hydrogen sulfide, was remarkable in providing protection to the plants against Cd toxicity.

Sub-lethal impacts of lead poisoning on blood biochemistry, immune function and delta-aminolevulinic acid dehydratase (δ-ALAD) activity in Cape (Gyps coprotheres) and white-backed (G. africanus) Vulture chicks

Although the prevalence of lead poisoning in southern Africa's Gyps vultures is now well-established, its finer physiological effects on these endangered species remain poorly characterised. We evaluated the sub-lethal impact of acute lead exposure on Cape and White-backed Vulture chicks from two breeding colonies in South Africa, by analysing its possible effects on key blood biochemistry parameters, immune function, packed cell volume and δ-aminolevulinic acid dehydratase (δ-ALAD) activity. All 37 White-backed Vulture nestlings sampled displayed elevated lead levels (>10 μg/dL), and seven had blood [Pb] >100 μg/dL. Eight of 28 Cape Vulture nestlings sampled had blood [Pb] exceeding background exposure, with one showing blood [Pb] >100 μg/dL. Delta-aminolevulinic acid dehydratase (δ-ALAD) activity was significantly and negatively related to blood [Pb] in nestlings from both species, with 50% inhibition of the enzyme predicted to occur at blood [Pb] = 52.8 μg/dL (White-backed Vulture) and 18.8 μg/dL (Cape Vulture). Although no significant relationship was found between % packed cell volume (PCV) and blood [Pb], the relatively lower mean PCV of 32.9% in White-backed Vulture chicks, combined with normal serum protein values, is likely indicative of depression or haemolytic anaemia. The leukogram was consistent in both species, although the presence of immature heterophils suggested an inflammatory response in White-backed Vulture chicks with blood [Pb] >100 μg/dL. Values for cholesterol, triglycerides, total serum protein, albumin, globulin, albumin/globulin ratio, alanine aminotransferase (ALT) and gamma-glutamyl transferase (GGT) were consistent with values previously reported. Calcium and phosphorus concentrations suggested no adverse effects on bone metabolism. A significant decrease in urea: uric acid (U:UA) ratio at blood [Pb] >100 μg/dL in White-backed Vulture chicks, brought about by a decrease in urea production, raises the possibility of hepatic abnormality. These results suggest that δ-ALAD activity may serve as a sensitive biomarker of lead toxicity in both species, while highlighting the need to better understand the significant variability in sensitivity that is observed, even between closely related members of the same genus.

Identification of common genes and pathways underlying imatinib and nilotinib treatment in CML: a Bioinformatics Study

Imatinib (IMA) and nilotinib are the first and second generations of BCR-ABL tyrosine kinase inhibitors, which widely applied in chronic myeloid leukemia (CML) treatment. Here we aimed to provide new targets for CML treatment by transcriptome analysis. Microarray data GSE19567 was downloaded and analyzed from Gene Expression Omnibus (GEO) to identify common genes, which are downregulated or upregulated in K562-imatinib and K562-nilotinib treated cells. The differentially expressed genes (DEGs) were assessed, and STRING and Cytoscape were used to create the protein–protein interaction (PPI) network. In imatinib and nilotinib treated groups’ comparison, there were common 626 upregulated and 268 downregulated genes, which were differentially expressed. The GO analysis represented the enrichment of DEGs in iron ion binding, protein tyrosine kinase activity, transcription factor activity, ATP binding, sequence-specific DNA binding, cytokine activity, the mitochondrion, sequence-specific DNA binding, plasma membrane and cell-cell adherens junction. KEGG pathway analysis revealed that downregulated DEGs were associated with pathways including microRNAs in cancer and PI3K-Akt signaling pathway. Furthermore, upregulated DEGs were involved in hematopoietic cell lineage, lysosome and chemical carcinogenesis. Among the upregulated genes, MYH9, MYH14, MYL10, MYL7, MYL5, RXRA, CYP1A1, FECH, AKR1C3, ALAD, CAT, CITED2, CPT1A, CYP3A5, CYP3A7, FABP1, HBD, HMBS and PPOX genes were found as hub genes. Moreover, 20 downregulated genes, YARS, AARS, SARS, GARS, CARS, IARS, RRP79, CEBPB, RRP12, UTP14A, PNO1, CCND1, DDX10, MYC, WDR43, CEBPG, DDIT3, VEGFA, PIM1 and TRIB3 were identified as hub genes. These genes have the potential to become target genes for diagnosis and therapy of CML patients.

HEPATO-PROTECTIVE ROLE OF DIPHENYL DISELENIDE IN STREPTOZOTOCININDUCED DIABETIC RATS

Diabetic hepatopathy prevalence as a result of prolonged hyperglycemia is increasing with the expanding size of the diabetes population around the world. In this present study, the hepato-protective activity of diphenyl diselenide (DPDSe) in a persistent diabetic state was studied. Diabetes mellitus were induced with streptozotocin (STZ) (60 mg/kg) intraperitoneally and after 60 days without treatment, the rats were administered diphenyl dislenide (DPDSe) (10 mg/kg) orally for 35 days. Animals were euthanized liver and serum were used for experimental determinations such as glycemic level, non enzymic and enzymic redox statuses, hepatic biomarkers, hematological and lipid prole. The results showed that the fasting glucose levels and thiobarbituric reactive substances were reduced at a signicant level of p &lt; 0.05. Furthermore, a concomitant rise in the percentage of DPPH free radicals scavenged, ferric reducing antioxidants property, protein and non-protein thiols were observed in hepatic tissue. The activities of delta–aminolevulinate dehydratase and antioxidant enzymes were improved at a signicant level of p &lt; 0.05. Additionally, there was signicant reduction (p &lt; 0.05) in the activity of the purinergic enzymes, hepatic biomarkers while there was an increase in the albumin level. Furthermore, the hematological and lipid prole of were refurbished. This study showed that DPDSe was able to reverse the biochemical alterations related with hepatic tissue in STZ-induced diabetic rats.

Characteristics of δ-Aminolevulenic Acid Dehydratase of the Cold-Water Sponge &lt;i&gt;Halisarca dujardinii&lt;/i&gt;

Delta-aminolevulinic acid dehydratase (ALAD) is a key enzyme in the cytoplasmic pathway of heme biosynthesis. Here, a primary structure of the ALAD gene of the marine cold-water sponge Halisarca dujardinii, a multimeric structure of the ALAD/hemB protein, and the ALAD gene expression during the sponge annual reproductive cycle were analyzed. On the base of results obtained one can suppose that the sponge ALAD gene expression is regulated by the transcription factor GATA-1 and DNA methylation. Re-aggregation of the sponge cells was accompanied by a decrease in ALAD expression and a change in the cellular content of the active ALAD/hemB form. Further study of heme biosynthesis and the role of ALAD/hemB in morphogenesis of basal animals may provide new opportunities for correcting pathologies in higher animals.

Assessment of heavy metal stress in the adaptation strategies of Tulipa luanica growing on serpentine soil through some biomarkers in comparison to Tulipa kosovarica

The aim of this study was to gain a better understanding of how Tulipa luanica adapts to growth in soil with higher concentrations of heavy metals and to assess potential toxic effects using various biomarkers, in comparison to Tulipa kosovarica, a typical serpentine species. For this purpose, we analyzed the concentrations of Al, Ca, Cd, Co, Cr, Cu, Fe, Mg, Mn, Ni, Pb, and Zn in the soil, as well as their accumulation in plants and their associated stress effects. The results indicate that, despite the presence of some metals in very high concentrations in the soil (Al, Fe, Mn, and Ni), they are translocated in minimal amounts within plant organs, particularly in T. luanica. Nearly all metals exhibited significantly higher concentrations in T. kosovarica when compared to T. luanica. Based on the analysis of biomarkers, it is apparent that T. luanica shows greater sensitivity to these conditions. This is evident through the decreased activity of δ-aminolevulinic acid dehydratase and levels of δ-aminolevulinic acid, malondialdehyde, and glutathione observed in T. luanica. It appears that T. luanica effectively restricts the absorption of metals in serpentine soils; however, it experiences oxidative stress induced by these metals, setting it apart from the more resilient T. kosovarica.

Expression deregulation of genes related to DNA repair and lead toxicity in occupationally exposed industrial workers.

Globally millions of people working in various industries and are exposed to different toxins which may affect their genetic stability and DNA integrity. Present study was designed to estimate the expression variation of genes related to DNA repair (XRCC1, PARP1) and lead toxicity (ALAD) in exposed industrial workers. About 200 blood samples were collected from workers of brick kiln, welding, furniture and paint industry (50/industry) along with age and gender matched controls. mRNA expression of genes was measured using RT-PCR. Serum levels of total ROS, POD, TBAR activity was calculated. Blood lead levels were estimated by atomic absorption spectrometer. Relative expression of XRCC1 and PARP1 gene was significantly (P < 0.001) upregulated, while ALAD gene expression was downregulated in exposed group compared to control. Expression of XRCC1 and PARP1 was increased (P < 0.001) in exposed workers with > 30year age compared to control with > 30year age. Same was observed when < 30year age group of control and exposed was compared. Likewise, XRCC1 and PARP1 expression was increased (P < 0.001) in exposed workers with > 30year age compared to workers with < 30year age. Whereas, ALAD gene showed significant (P < 0.01) decrease in > 30year age workers compared to control of same age and exposed with < 30year of age. Relative expression of XRCC1 and PARP1 was increased (P < 0.001) inexposed smokers compared to exposed non-smokers and control smokers. Whereas, ALAD gene expression reduced (P < 0.001) significantly in both groups. Blood lead content was higher (P < 0.001) in exposed group compared to control. Strong correlation was observed between XRCC1, PARP1 and ALAD gene versus age, total exposure duration, exposure per day and lead deposition. ROS, TBARS and POD activity was higher (P < 0.01) in exposed group compared to control group. Present study suggested deregulation of genes related to DNA repair and lead intoxication in exposed group compared to controls. Strong correlation was observed between selected genes and demographic parameters. Present results revealed altered activity of oxidative stress markers which would induce oxidative damage to DNA integrity and limit the function of repair enzymes.