Caulerpa chemnitzia polysaccharide exerts immunomodulatory activity in macrophages by mediating the succinate/PHD2/HIF-1α/IL-1ß pathway.

Algal polysaccharide is an important food functional factor with diverse bioactive and low toxicity. Previous studies have confirmed Caulerpa chemnitzia polysaccharides (CRVP) have immunomodulatory activity, but the immunomodulatory mechanism of CRVP in macrophages has not been thoroughly explored yet. In our research, we found that CRVP has outstanding immunomodulatory activity in macrophages, which is reflected in promoting cell proliferation, upregulating cytokines (IL-1ß, IL-6, and TNF-α) expression, and increasing NO and ROS levels. Additionally, the result of joint analysis of untargeted metabolomics showed metabolism played a major role in the immunomodulatory of CRVP and suggested succinic acid was a key metabolite. Further verification indicated that the accumulation of succinic acid in macrophages after administered with CRVP, induced the down-regulation of prolyl hydroxylase domain 2 (PHD2) and up-regulation of hypoxia-inducible factor-1α (HIF-1α), thereby enhancing IL-1ß expression. Together, the immunomodulatory activity of CRVP in macrophages via succinate/PHD2/HIF-1α/IL-1ß pathway.

Succinate level is increased and succinate dehydrogenase exerts forward and reverse catalytic activities in lipopolysaccharides-stimulated cardiac tissue: The protective role of dimethyl malonate.

This study aimed to investigate the alterations of myocardial succinate and fumarate levels with or without succinate dehydrogenase (SDH) inhibitor dimethyl malonate during 24 h of lipopolysaccharides (LPS) challenge, as well as the effects of dimethyl malonate on the impaired cardiac tissue. Myocardial succinate and fumarate levels were increased in the initial 9 h of LPS challenge. During this time, dimethyl malonate increased the succinate level, decreased the fumarate level, aggravated the cardiac dysfunction, reduced the oxidative stress, had little effect on interleukin-1ß production, promoted interleukin-10 production and bothered the ATP production. Co-treatment with exogenous succinate significantly increased interleukin-1ß production in this period. After 12 h of LPS challenge, myocardial the succinate level increased sharply, while the fumarate level gradually decreased. During 12-24 h of LPS challenge, dimethyl malonate effectively reduced the succinate level, increased the fumarate level, improved cardiac dysfunction, inhibited interleukin-1ß production, and had little effect on oxidative stress, interleukin-10 production, and ATP production. LPS challenge also significantly increased the myocardial succinate receptor 1 expression and circulating succinate level. Inhibition of succinate receptor 1 significantly reduced the mRNA expression of interleukin-1ß. In conclusion, the current study suggests that myocardial succinate accumulates during LPS challenge, and that SDH activity may be transformed (from forward to reversed) and involved in a line of stress response. Dimethyl malonate inhibits SDH and, depending on the time of treatment, reduces LPS-induced cardiac impairment. Furthermore, accumulated succinate exerts pro-inflammatory effects partly via succinate receptor 1 signaling.

Multi-Omics Analysis Revealed a Significant Alteration of Critical Metabolic Pathways Due to Sorafenib-Resistance in Hep3B Cell Lines.

Hepatocellular carcinoma (HCC) is the second prominent cause of cancer-associated death worldwide. Usually, HCC is diagnosed in advanced stages, wherein sorafenib, a multiple target tyrosine kinase inhibitor, is used as the first line of treatment. Unfortunately, resistance to sorafenib is usually encountered within six months of treatment. Therefore, there is a critical need to identify the underlying reasons for drug resistance. In the present study, we investigated the proteomic and metabolomics alterations accompanying sorafenib resistance in hepatocellular carcinoma Hep3B cells by employing ultra-high-performance liquid chromatography quadrupole time of flight mass spectrometry (UHPLC-QTOF-MS). The Bruker Human Metabolome Database (HMDB) library was used to identify the differentially abundant metabolites through MetaboScape 4.0 software (Bruker). For protein annotation and identification, the Uniprot proteome for Homo sapiens (Human) database was utilized through MaxQuant. The results revealed that 27 metabolites and 18 proteins were significantly dysregulated due to sorafenib resistance in Hep3B cells compared to the parental phenotype. D-alanine, L-proline, o-tyrosine, succinic acid and phosphatidylcholine (PC, 16:0/16:0) were among the significantly altered metabolites. Ubiquitin carboxyl-terminal hydrolase isozyme L1, mitochondrial superoxide dismutase, UDP-glucose-6-dehydrogenase, sorbitol dehydrogenase and calpain small subunit 1 were among the significantly altered proteins. The findings revealed that resistant Hep3B cells demonstrated significant alterations in amino acid and nucleotide metabolic pathways, energy production pathways and other pathways related to cancer aggressiveness, such as migration, proliferation and drug-resistance. Joint pathway enrichment analysis unveiled unique pathways, including the antifolate resistance pathway and other important pathways that maintain cancer cells' survival, growth, and proliferation. Collectively, the results identified potential biomarkers for sorafenib-resistant HCC and gave insights into their role in chemotherapeutic drug resistance, cancer initiation, progression and aggressiveness, which may contribute to better prognosis and chemotherapeutic outcomes.

[Succinate salts in solving the «oxygen paradox¼ of reperfusion]. / Suktsinaty v reshenii «kislorodnogo paradoksa¼ reperfuzii.

Reperfusion damage to the cellular structures of tissues in the early post-ischemic period is a consequence of the restoration of blood flow and reoxygenation. Currently, there is no effective treatment for reperfusion metabolic disorders in clinical practice. Over the past decades, biological studies of hypoxia and the role of hypoxia-inducible factor-1α (HIF-1), potentiating succinatoxidase oxidation by signal from the succinate-dependent receptor (GPR91), have significantly improved the understanding of oxygen homeostasis during the period of recovery of blood flow. HIF-1 plays a key role in postischemic damage and is an oxygen-sensitive transcription factor that mediates adaptive metabolic responses to hypoxia and hyperoxia during reperfusion and reoxygenation. Activation of HIF-1 by succinate improves cell survival in hypoxic and posthypoxic (hyperoxygenated) environment, altering energy metabolism, proliferation, angiogenesis and vascular remodeling. The role of succinate oxidation in the period of ischemia / reperfusion and reoxygenation suggests the widespread use of infusion succinate as a protector that reduces the degree of tissue damage by reactive oxygen species (ROS) and restores the usual oxygen homeostasis.

Lack of Major Genome-Wide DNA Methylation Changes in Succinate-Treated Human Epithelial Cells.

The tricarboxylic acid (TCA) metabolite, succinate, is a competitive inhibitor of dioxygenase enzymes that require alpha ketoglutarate as a cofactor. One family of dioxygenases are the ten-eleven translocation (TET) proteins, which oxidize 5-methylcytosine to promote DNA demethylation. Inhibition of DNA demethylation is expected to lead to DNA hypermethylation, at least at genomic regions at which TET proteins are engaged. We treated human bronchial epithelial cells with succinate for five days and confirmed its effect on TET protein function by observing diminished formation of 5-hydroxymethylcytosine, the first oxidation product of the TET enzymatic reaction. We then analyzed global DNA methylation patterns by performing whole-genome bisulfite sequencing. Unexpectedly, we did not observe differentially methylated regions (DMRs) that reached genome-wide statistical significance. We observed a few regions of clustered DNA hypomethylation, which was also not expected based on the proposed mechanisms. We discuss potential explanations for our observations and the implications of these findings for tumorigenesis.

Low-concentration iron promotes Klebsiella pneumoniae biofilm formation by suppressing succinic acid.

BACKGROUND: Klebsiella pneumoniae is widely distributed in water and plays a major role in both human and poultry infections. Many K. pneumoniae strains form biofilms on various surfaces, enhancing their pathogenicity and resistance to antibiotics. The water supply pipeline of chicken farms has become a hotbed for the growth of K pneumoniae biofilm because of its humid environment, and because the chicken drinking water pipeline is thin, it is easily blocked by the biofilm, and the diffused cells can cause repeated and persistent infections. Iron is vital to the growth of microorganisms and the formation of biofilms. Therefore, the aim of this study was to examine the effects of iron on K. pneumoniae biofilm formation and any associated metabolic changes to provide a rationale for reducing the formation of biofilms. RESULTS: Biofilm formation was enhanced to the greatest extent by the presence of 0.16 mM FeCl2, producing a denser structure under electron microscopy. The number of biofilm-forming and planktonic bacteria did not change, but protein and polysaccharide concentrations in the bacterial extracellular polymeric substances (EPS) were significantly increased by iron supplementation. To clarify this mechanism, intracellular metabolomic analysis was carried out, showing that the differential, down-regulated metabolites included succinic acid. The addition of 1.7 mM succinic acid counteracted the biofilm-forming effect of iron, with no bactericidal side effects. CONCLUSION: This study demonstrates the importance of succinic acid and iron in K. pneumoniae biofilms, and provides insight into the formation of K. pneumoniae biofilms and direction for the development of new antibacterial agents.

The effect of pesticides on the NADH-supported mitochondrial respiration of permeabilized potato mitochondria.

Pesticides can seriously affect the respiratory chain of the mitochondria of many crops, reducing the intensity of plant growth and its yield. Studying the effect of pesticides on the bioenergetic parameters of intact plant mitochondria is a promising approach for assessing their toxicity. In this study, we investigated the effect of some pesticides on isolated potato mitochondria, which used exogenous NADH as a substrate for respiration. We showed that succinate is the most preferred substrate for phosphorylating respiration of intact potato tubers mitochondria. Potato mitochondria poorly oxidize exogenous NADH, despite of the presence of external NADH dehydrogenases. Permeabilization of the mitochondrial membrane with alamethicin increased the availability of exogenous NADH to complex I. However, the pathway of electrons through complex I to complex IV makes intact potato mitochondria susceptible to a number of pesticides such as difenoconazole, fenazaquin, pyridaben and tolfenpyrad, which strongly inhibit the rate of mitochondrial respiration. However, these pesticides only slightly inhibited the rate of oxygen consumption during succinate-supported respiration. Dithianon, the inhibitor of Complex II, is the only pesticide which significantly increased the respiratory rate of NADH-supported respiration of permeabilized mitochondria of potato. Thus, it can be assumed that the alternative NADH dehydrogenases for electron flow represent a factor responsible for plant resistance to xenobiotics, such as mitochondria-targeted pesticides.

Intestinal epithelial tuft cell induction is negated by a murine helminth and its secreted products.

Helminth parasites are adept manipulators of the immune system, using multiple strategies to evade the host type 2 response. In the intestinal niche, the epithelium is crucial for initiating type 2 immunity via tuft cells, which together with goblet cells expand dramatically in response to the type 2 cytokines IL-4 and IL-13. However, it is not known whether helminths modulate these epithelial cell populations. In vitro, using small intestinal organoids, we found that excretory/secretory products (HpES) from Heligmosomoides polygyrus blocked the effects of IL-4/13, inhibiting tuft and goblet cell gene expression and expansion, and inducing spheroid growth characteristic of fetal epithelium and homeostatic repair. Similar outcomes were seen in organoids exposed to parasite larvae. In vivo, H. polygyrus infection inhibited tuft cell responses to heterologous Nippostrongylus brasiliensis infection or succinate, and HpES also reduced succinate-stimulated tuft cell expansion. Our results demonstrate that helminth parasites reshape their intestinal environment in a novel strategy for undermining the host protective response.

Cell-Permeable Succinate Rescues Mitochondrial Respiration in Cellular Models of Amiodarone Toxicity.

Amiodarone is a potent antiarrhythmic drug and displays substantial liver toxicity in humans. It has previously been demonstrated that amiodarone and its metabolite (desethylamiodarone, DEA) can inhibit mitochondrial function, particularly complexes I (CI) and II (CII) of the electron transport system in various animal tissues and cell types. The present study, performed in human peripheral blood cells, and one liver-derived human cell line, is primarily aimed at assessing the concentration-dependent effects of these drugs on mitochondrial function (respiration and cellular ATP levels). Furthermore, we explore the efficacy of a novel cell-permeable succinate prodrug in alleviating the drug-induced acute mitochondrial dysfunction. Amiodarone and DEA elicit a concentration-dependent impairment of mitochondrial respiration in both intact and permeabilized platelets via the inhibition of both CI- and CII-supported respiration. The inhibitory effect seen in human platelets is also confirmed in mononuclear cells (PBMCs) and HepG2 cells. Additionally, amiodarone elicits a severe concentration-dependent ATP depletion in PBMCs, which cannot be explained solely by mitochondrial inhibition. The succinate prodrug NV118 alleviates the respiratory deficit in platelets and HepG2 cells acutely exposed to amiodarone. In conclusion, amiodarone severely inhibits metabolism in primary human mitochondria, which can be counteracted by increasing mitochondrial function using intracellular delivery of succinate.

The apoptotic efficacy of succinic acid on renal cancer cell lines.

Recently, studies on the effects of non-toxic substances on cancer prophylaxis have gained value as an alternative to existing treatment options. Current studies have shown that succinic acid or its derivatives exhibit anticancer activity by inducing apoptosis. We aimed to investigate the anticancer activity of succinic acid on renal cancer for the first time in the literature. The cytotoxic activity of succinic acid on CAKI-2 and ACHN as renal cancer cell lines and MRC-5 as a healthy cell line was determined using the WST-1 cytotoxicity test. Apoptotic activity was measured by Annexin V test and cell death ELISA kit. The results showed that 25 µM and 50 µM doses of succinic acid for 24 h remarkably reduced the cell viability for CAKI-2 cells (89.77% and 90.77%) and ACHN cells (41.57% and 54.54%). Also, no significant effect was observed on the healthy cell line, as we expected. Additionally, administration of succinic acid at same doses resulted in apoptotic activity for ACHN cells (19.1 and 12.7) and CAKI-2 cells (19.85 and 29.55). ELISA results with same doses of succinic acid treatment increased the apoptotic fragment rates by 4.7 and 2.13-fold in CAKI-2 cells, and 32.92, 12.7-fold in ACHN cells. Succinic acid is a focal point for cancer treatments not only for its apoptotic success on cancer cells but also for its capacity to be metabolically active for humans. Our results suggest that succinic acid could be a potential therapeutic agent for individual cancer treatment approaches together with further molecular research.

Poly (Lactic-co-Glycolic Acid) & Tocopheryl Polyethylene Glycol Succinate Nanoparticles for the Treatment of Different Brain Cancers.

Nanotechnology and material science developments emerge in the manufacturing of various novel modes of drug delivery, which have proven scientifically promising. Polymer nanoparticles have high stability, high specificity, high drug-carrying power, control release, and potential to be used in various pathways. They usually supply hydrophilic and hydrophobic molecules with medicines. In this review, we have discussed the different types of brain tumour, different PLGA (Poly Lactic-co-Glycolic Acid) nanostructures, PLGA in brain tumour targeting, and the recent advancement of PLGA based nanoparticles. This review focused on the method of preparation of polymeric nanoparticles, the significance of EPR (Enhanced Permeability and Retention) effect with PLGA, the significance of TPGS in cancer, and discussed the pharmaceutical application of PLGA nanoparticles. We expect these polymeric nanoparticles will be very successful and efficient for disease targeting in the future and new techniques will emerge.

Mitoflash generated at the Qo site of mitochondrial Complex III.

The previous research has shown that mitochondrial flash (mitoflash) genesis are functionally and mechanistically integrated with mitochondrial electron transport chain (ETC) energy metabolism. However, the response of mitoflash to superoxide is not entirely consistent with the response of MitoSOX Red. The generation mechanism of mitoflash is still unclear. Here, we investigated mitoflash activities, using the different combinations of ETC substrates and inhibitors, in permeabilized cardiomyocytes or hearts. We found that blocking the complete electron flow, from Complex I to IV, with any one of ETC inhibitors including rotenone (Rot), antimycin A (AntA), myxothiazol (Myxo), stigmatellin, and sodium cyanide, will lead to the abolishment of mitoflashes triggered by substrates in adult permeabilized cardiomyocytes. However, Myxo boosted mitoflashes triggered by the reverse electron of N,N,N',N'-tetramethyl-p-phenylenediamine/ascorbate. Moreover, Rot and AntA furtherly enhanced mitoflash activity rather than depressed it, suggesting that mitoflashes generated at the Complex III Qo site. Meanwhile, the inhibition of Complex III protein expression resulted in the activity of Complex III decrease, which decreased mitoflash frequency. The function defect (no change of protein level) of the Qo site of Complex III in aging hearts augmented mitoflash generation confirmed the Qo site function was critical to mitoflash genesis. Thus, our results indicate that mitoflash detected by circularly permuted yellow fluorescent protein is generated at the Qo site of Complex III.

In Vitro Inhibitory Effect of Succinic Acid on T-Cell Acute Lymphoblastic Leukemia Cell Lines.

BACKGROUND AND AIMS: Although several treatment regimens for T-cell acute lymphoblastic leukemia (T-ALL), trouble is still ongoing that relapse of disease after therapies in both pediatric and adult patients. Hence, the demand for new alternative therapeutics that are antiproliferative for cancer cells but do not harm healthy cells in treatments is increasing day by day. This study aimed to investigate whether succinic acid show anti-proliferative and apoptotic effect of on T-ALL cell lines. METHODS: Time and dose-dependent effects of succinic acid on T-ALL cell lines were determined by using WST-1, caspase-3/ bicinchoninic acid (BCA), and Annexin V-Fluorescein isothiocyanate (FITC) assays. We included the MRC-5 cell line in our study as a healthy control group. RESULTS: Based on our findings, 25 and 50 mmol dosages of succinic acid has shown an apoptotic effect on T-ALL cell lines for 48 h treatment. Also, it has shown that after 48 h exposure of 25 and 50 mmol dosages of succinic acid has no significant cytotoxic effect in healthy MRC-5 cells. Apoptotic activity of succinic acid on CCRF-CEM cell line was caspase-3 dependent but not for MOLT-4. As a consequence, succinic acid was found to effect for T-ALL treatment in vitro and might also enlighten new study fields for different cancer experiments.

Receptor GPR91 contributes to voiding function and detrusor relaxation mediated by succinate.

AIM: Succinate activates the receptor GPR91 identified in the bladder. The present study aims to unravel the mechanisms of bladder relaxation by succinate and how the receptor is involved in structural and functional changes of the bladder. METHODS: Physiological recordings of bladder function were carried out by cystometry and organ bath from C57BL/6 mice, homozygous GPR91-/- mice, and Sprague-Dawley (SD) rats. GPR91 expression was confirmed by polymerase chain reaction and tissue morphology was examined by light (Masson trichrome) and fluorescence microscopy. Nitric oxide (NO) and ATP secretion were measured. RESULTS: Bladders of GPR91 KO mice had a greater mass to body weight ratio with a thicker bladder wall compared to C57BL/6 mice. They also displayed increased basal and maximal bladder pressures, and decreased intercontraction intervals, bladder capacity, micturition volume, and compliance. During cystometry, bladders of SD rats and C57BL/6 mice instilled with succinate (10 mM) showed signs of relaxation while bladders of GPR91 KO mice were unresponsive. Similarly, in organ bath, succinate relaxed bladder strips preincubated with carbachol, except GPR91 KO ones. Relaxation was stronger in the presence of urothelium and independent of NO synthesis. Bladder strips from all mice groups showed similar responses to KCl, carbachol, and electrical stimulation. In vitro, succinate increased NO secretion in urothelial cell culture of both C57BL6 and GPR91 KO mice while ATP secretion was potently decreased by succinate in C57BL6 culture only. CONCLUSION: Succinate through GPR91 is essential to bladder structure and contraction. GPR91 relaxes the detrusor partially by decreasing urothelial ATP secretion.

Glycogen phosphorylase inhibitor, 2,3-bis[(2E)-3-(4-hydroxyphenyl)prop-2-enamido] butanedioic acid (BF142), improves baseline insulin secretion of MIN6 insulinoma cells.

Type 2 diabetes mellitus (T2DM), one of the most common metabolic diseases, is characterized by insulin resistance and inadequate insulin secretion of ß cells. Glycogen phosphorylase (GP) is the key enzyme in glycogen breakdown, and contributes to hepatic glucose production during fasting or during insulin resistance. Pharmacological GP inhibitors are potential glucose lowering agents, which may be used in T2DM therapy. A natural product isolated from the cultured broth of the fungal strain No. 138354, called 2,3-bis(4-hydroxycinnamoyloxy)glutaric acid (FR258900), was discovered a decade ago. In vivo studies showed that FR258900 significantly reduced blood glucose levels in diabetic mice. We previously showed that GP inhibitors can potently enhance the function of ß cells. The purpose of this study was to assess whether an analogue of FR258900 can influence ß cell function. BF142 (Meso-Dimethyl 2,3-bis[(E)-3-(4-acetoxyphenyl)prop-2-enamido]butanedioate) treatment activated the glucose-stimulated insulin secretion pathway, as indicated by enhanced glycolysis, increased mitochondrial oxidation, significantly increased ATP production, and elevated calcium influx in MIN6 cells. Furthermore, BF142 induced mTORC1-specific phosphorylation of S6K, increased levels of PDX1 and insulin protein, and increased insulin secretion. Our data suggest that BF142 can influence ß cell function and can support the insulin producing ability of ß cells.

Succinate Activates EMT in Intestinal Epithelial Cells through SUCNR1: A Novel Protagonist in Fistula Development.

The pathogenesis of Crohn's disease-associated fibrostenosis and fistulas imply the epithelial-to-mesenchymal transition (EMT) process. As succinate and its receptor (SUCNR1) are involved in intestinal inflammation and fibrosis, we investigated their relevance in EMT and Crohn's disease (CD) fistulas. Succinate levels and SUCNR1-expression were analyzed in intestinal resections from non-Inflammatory Bowel Disease (non-IBD) subjects and CD patients with stenosing-B2 or penetrating-B3 complications and in a murine heterotopic-transplant model of intestinal fibrosis. EMT, as increased expression of Snail1, Snail2 and vimentin and reduction in E-cadherin, was analyzed in tissues and succinate-treated HT29 cells. The role played by SUCNR1 was studied by silencing its gene. Succinate levels and SUCNR1 expression are increased in B3-CD patients and correlate with EMT markers. SUCNR1 is detected in transitional cells lining the fistula tract and in surrounding mesenchymal cells. Grafts from wild type (WT) mice present increased succinate levels, SUCNR1 up-regulation and EMT activation, effects not observed in SUCNR1-/- tissues. SUCNR1 activation induces the expression of Wnt ligands, activates WNT signaling and induces a WNT-mediated EMT in HT29 cells. In conclusion, succinate and its receptor are up-regulated around CD-fistulas and activate Wnt signaling and EMT in intestinal epithelial cells. These results point to SUCNR1 as a novel pharmacological target for fistula prevention.

H2S-stimulated bioenergetics in chicken erythrocytes and the underlying mechanism.

The production of H2S and its effect on bioenergetics in mammalian cells may be evolutionarily preserved. Erythrocytes of birds, but not those of mammals, have a nucleus and mitochondria. In the present study, we report the endogenous production of H2S in chicken erythrocytes, which was mainly catalyzed by 3-mercaptopyruvate sulfur transferase (MST). ATP content of erythrocytes was increased by MST-generated endogenous H2S under normoxic, but not hypoxic, conditions. NaHS, a H2S salt, increased ATP content under normoxic, but not hypoxic, conditions. ATP contents in the absence or presence of NaHS were eliminated by different inhibitors for mitochondrial electron transport chain in chicken erythrocytes. Succinate and glutamine, but not glucose, increased ATP content. NaHS treatment similarly increased ATP content in the presence of glucose, glutamine, or succinate, respectively. Furthermore, the expression and activity of sulfide:quinone oxidoreductase were enhanced by NaHS. The structural integrity of chicken erythrocytes was largely maintained during 2-wk NaHS treatment in vitro, whereas most of the erythrocytes without NaHS treatment were lysed. In conclusion, H2S may regulate cellular bioenergetics as well as cell survival of chicken erythrocytes, in which the functionality of the electron transport chain is involved. H2S may have different regulatory roles and mechanisms in bioenergetics of mammalian and bird cells.

Rhizosphere assisted biodegradation of benzo(a)pyrene by cadmium resistant plant-probiotic Serratia marcescens S2I7, and its genomic traits.

Melia azedarach-rhizosphere mediated degradation of benzo(a)pyrene (BaP), in the presence of cadmium (Cd) was studied, using efficient rhizobacterial isolate. Serratia marcescens S2I7, isolated from the petroleum-contaminated site, was able to tolerate up to 3.25 mM Cd. In the presence of Cd, the isolate S2I7 exhibited an increase in the activity of stress-responsive enzyme, glutathione-S-transferase. Gas Chromatography-Mass spectroscopy analysis revealed up to 59% in -vitro degradation of BaP after 21 days, while in the presence of Cd, the degradation was decreased by 14%. The bacterial isolate showed excellent plant growth-promoting attributes and could enhance the growth of host plant in Cd contaminated soil. The 52,41,555 bp genome of isolate S. marcescens S2I7 was sequenced, assembled and annotated into 4694 genes. Among these, 89 genes were identified for the metabolism of aromatic compounds and 172 genes for metal resistance, including the efflux pump system. A 2 MB segment of the genome was identified to contain operons for protocatechuate degradation, catechol degradation, benzoate degradation, and an IclR type regulatory protein pcaR, reported to be involved in the regulation of protocatechuate degradation. A pot trial was performed to validate the ability of S2I7 for rhizodegradation of BaP when applied through Melia azedarach rhizosphere. The rhizodegradation of BaP was significantly higher when augmented with S2I7 (85%) than degradation in bulk soil (68%), but decreased in the presence of Cd (71%).

Acute lymphoblastic leukaemia patients treated with PEGasparaginase develop antibodies to PEG and the succinate linker.

Polyethylene glycol (PEG) conjugated asparaginase (PEGasparaginase) is essential for treatment of paediatric acute lymphoblastic leukaemia. We developed an assay identifying antibodies against the PEG-moiety, the linker and the drug itself in patients experiencing hypersensitivity reactions to PEGasparaginase. Eighteen patients treated according to the DCOG ALL-11 protocol, with a neutralizing hypersensitivity reaction to PEGasparaginase to the first PEGasparaginase doses in induction (12 patients) or during intensification after interruption of several months (6 patients) were included. ELISA was used to measure antibodies, coating with the succinimidyl succinate linker conjugated to BSA, PEGfilgrastim and Escherichia coli asparaginase, and using hydrolysed PEGasparaginase and mPEG5,000 for competition. Anti-PEG antibodies were detected in all patients (IgG 100%; IgM 67%) of whom 39% had anti-PEG antibodies exclusively. Pre-existing anti-PEG antibodies were also detected in patients who not previously received a PEGylated therapeutic (58% IgG; 21% IgM). Antibodies against the SS-linker were predominantly detected during induction (50% IgG; 42% IgM). Anti-asparaginase antibodies were detected in only 11% during induction but 94% during intensification. In conclusion, anti-PEG and anti-SS-linker antibodies predominantly play a role in the immunogenic response to PEGasparaginase during induction. Thus, switching to native E. coli asparaginase would be an option for adequate asparaginase treatment.

Mitochondrial Ca2+ -activated F1 FO -ATPase hydrolyzes ATP and promotes the permeability transition pore.

The properties of the mitochondrial F1 FO -ATPase catalytic site, which can bind Mg2+ , Mn2+ , or Ca2+ and hydrolyze ATP, were explored by inhibition kinetic analyses to cast light on the Ca2+ -activated F1 FO -ATPase connection with the permeability transition pore (PTP) that initiates cascade events leading to cell death. While the natural cofactor Mg2+ activates the F1 FO -ATPase in competition with Mn2+ , Ca2+ is a noncompetitive inhibitor in the presence of Mg2+ . Selective F1 inhibitors (Is-F1 ), namely NBD-Cl, piceatannol, resveratrol, and quercetin, exerted different mechanisms (mixed and uncompetitive inhibition) on either Ca2+ - or Mg2+ -activated F1 FO -ATPase, consistent with the conclusion that the catalytic mechanism changes when Mg2+ is replaced by Ca2+ . In a partially purified F1 domain preparation, Ca2+ -activated F1 -ATPase maintained Is-F1 sensitivity, and enzyme inhibition was accompanied by the maintenance of the mitochondrial calcium retention capacity and membrane potential. The data strengthen the structural relationship between Ca2+ -activated F1 FO -ATPase and the PTP, and, in turn, on consequences, such as physiopathological cellular changes.