Polymorphisms and haplotypes of serine hydroxymethyltransferase and risk of squamous cell carcinoma of the head and neck: a case-control analysis

Low dietary intake of fruits and vegetables, particularly folate deficiency, has been associated with the risk of squamous cell carcinoma of the head and neck (SCCHN). We hypothesized that polymorphisms of the cytosolic serine hydroxymethyltransferase (SHMT1) gene involved in folate-dependent, one-carbon metabolism are associated with SCCHN risk. In a hospital-based, case-control study of 721 non-Hispanic white SCCHN patients and 1,234 control subjects, frequency-matched by age and sex, three known SHMT1 polymorphisms (34761C>T, 34840C>G and 34859C>T) were genotyped. It was found that none of these three polymorphisms alone had a significant main effect on the risk of SCCHN. However, when the three polymorphisms were evaluated together by the number of the variant (risk) haplotype alleles (i.e. 34761 T, 34840G or 34859 T), the risk of SCCHN was significantly increased in a dose-response manner as the number of variant haplotype alleles increased compared to those with zero variant alleles [adjusted odd ratio (OR)=1.39, 95% confidence interval (CI)=1.14-1.70 for 1-3 variant alleles and OR=1.46, 95% CI=1.09-1.97 for 4-6 variant alleles; Ptrend=0.001]. In stratification analysis, this significant association was confined to younger men (相似文献   

The reduced folate carrier gene is a novel selectable marker for recombinant protein overexpression

Folate cofactors are one-carbon donors essential for the biosynthesis of purines and thymidylate. Mammalian cells are devoid of folate biosynthesis and are therefore folate auxotrophs that take up folate vitamins primarily via the reduced folate carrier (RFC). In this study, we showed that the human RFC (hRFC) gene can serve as a novel selectable marker for the overproduction of recombinant proteins. Toward this end, a hemagglutinin (HA) epitope tagged hRFC (hRFC-HA) was introduced into a bicistronic vector (pIRES2-EGFP), upstream of an enhanced green fluorescent protein (EGFP) reporter gene. Chinese hamster ovary cells deficient in RFC activity were isolated and transfected with this construct, followed by gradual deprivation of leucovorin, the sole folate source in the growth medium. Only cells with hRFC-HA overexpression were able to take up leucovorin and thereby survive these selective conditions. Western blot and immunofluorescence analyses confirmed that the hRFC-HA was overexpressed at extremely high levels, properly glycosylated and sorted out to the plasma membrane. This resulted in a approximately 450-fold increase in [3H]methotrexate influx and approximately 100-fold increased sensitivity to methotrexate, relative to untransfected RFC-deficient cells. Flow cytometric analysis consistently revealed that EGFP was overexpressed approximately 100-fold above the autofluorescence level. Overproduction of hRFC-HA and EGFP was stably maintained for at least 2 months in a constant concentration of leucovorin. These results establish a novel RFC-based metabolic selection system for the efficient overexpression of recombinant proteins. Furthermore, the possible implications to subcellular transporter localization and restoration of MTX sensitivity in drug-resistant tumors by RFC-based gene therapy are discussed.     

Impact of SHMT1 polymorphism on the clinical outcome of patients with metastatic colorectal cancer treated with first-line FOLFIRI+bevacizumab

The impact of thymidylate synthase (TYMS), methylenetetrahydrofolate reductase (MTHFR), and serine hydroxymethyltransferase 1 (SHMT1) gene polymorphisms and that of dihydropyrimidine dehydrogenase (DPD) enzyme activity, serum total homocysteine level, and estimated serum creatinine clearance on first-line 5-fluorouracil, leucovorin, irinotecan, and bevacizumab (FOLFIRI+bevacizumab) regimen efficacy in metastatic colorectal cancer patients was investigated. DNA was extracted from peripheral blood mononuclear cells. Genotyping was performed for TYMS 5'UTR variable number tandem repeat, TYMS 3'UTR ins/del, MTHFR C677T, and SHMT1 C1420T polymorphisms. The DPD activity of peripheral blood mononuclear cells was also determined. The univariate and multivariate analyses demonstrated that the SHMT1 1420T allele was associated with better response (P=0.025) and longer progression-free survival (PFS) (P=0.00004) and overall survival (OS) (P=0.034). Grade ≥2 hypertension was also an independent prognostic factor of longer progression-free survival and OS. Bevacizumab-related hypertension might be a predictive marker of treatment efficacy (P=0.0002 for OS) in the case of wild (CC) SHMT1 1420 genotypes only.     

Suspected methotrexate toxicity from omeprazole: a case review of carboxypeptidase G2 use in a methotrexate-experienced patient with methotrexate toxicity and a review of the literature

We report a case of methotrexate toxicity potentially induced by a drug interaction between methotrexate and omeprazole in a 25-year-old man with osteosarcoma. The patient was placed on omeprazole after his first cycle of high-dose methotrexate for stress ulcer prophylaxis, and it was discontinued before the start of the first day of the patient's second round of high-dose methotrexate. The 24-hour methotrexate level was elevated and he continued to have sustained levels for 18 days. Side effects due to elevated serum methotrexate included seizures, mucositis, acute renal failure, and thrombocytopenia. Aggressive hydration, urinary alkalinization, and leucovorin were continued during the period of elevated methotrexate levels, with the patient receiving a course of hemodialysis and a dose of carboxypeptidase G2. The patient's symptoms resolved, and his renal function returned to baseline within 2 months. The patient was able to receive future courses of chemotherapy without methotrexate. Although use of the Naranjo adverse reaction probability scale indicated a probable relationship (score of 6) between the patient's development of methotrexate toxicity and omeprazole use, we believe this was a drug-drug interaction case consistent with previous reports in the literature.     

Evidence of competitive inhibition of methotrexate absorption by leucovorin calcium in rat small intestine

The effect of leucovorin calcium on the intestinal absorption of methotrexate in rat small intestine was investigated using an in situ rat gut technique. First, the kinetic absorption in situ parameters for methotrexate in solution were obtained: V_m=21.54 (±2.22) μM/h; K_m=10.51 (±1.08) μM; k_a=0.26 (±0.03) h⁻¹ and AIC=−188.63. The inhibitory effect of leucovorin calcium in methotrexate intestinal absorption has been investigated by perfusing of 10 μM methotrexate isotonic solutions containing increasing concentrations of leucovorin calcium (10–500 μM), and the remaining concentrations of both compounds were measured. A competitive inhibition of methotrexate absorption was detected: the apparent absorption rate constant of the drug decreased as the initial leucovorin calcium concentration increased. Higher leucovorin calcium concentrations, however, did not completely abolish the absorption of the drug (at 500 μM of leucovorin calcium, only 84% inhibition was observed). Apparent parameters characterizing the absorption of leucovorin calcium in the presence of methotrexate 10 μM were: V_mi=14.70 (±1.74) μM; K_mi=9.43 (±1.59) μM; k_ai=0.28 (±0.02) h⁻¹; AIC=−191.53). We can concluded that methotrexate and leucovorin calcium compete for the same intestinal carrier system. This means that since leucovorin calcium, because of its ready conversion to other tetrahydrofolic derivatives (McEvoy, 1996. AHFS Drug Information, Bethesda, MD, pp. 751–758), is administered together with methotrexate in order to prevent the hematopoietic and reticuloendothelial toxic effects of folic acid antagonists, using high leucovorin calcium concentrations, when the urine excretion is decreased, could prevent intestinal drug reabsorption and the drug could then be excreted in the feces, thereby decreasing the risk of poisoning.     

High-dose methotrexate in acute lymphoblastic leukemia: where is the evidence for its continued use?

High-dose intravenous methotrexate is an important component of many effective chemotherapeutic regimens for childhood acute lymphoblastic leukemia (ALL). Its use has a strong pharmacologic rationale: to overcome mechanisms of resistance of the malignant cells and to achieve cytotoxic concentrations in sanctuary sites for lymphoblasts. Although therapeutic progress in ALL during the past 4 decades has been closely associated with more widespread use of intravenous methotrexate and in progressively larger doses, little data exist to clearly support the use of high-dose intravenous methotrexate over a regimen of prolonged administration of low-dose methotrexate. The implied superiority of intravenous methotrexate mainly stems from studies that used identical leucovorin rescue with low-dose methotrexate or from studies of upfront window therapy in untreated children with ALL in which single standard doses of oral methotrexate were compared with high-dose intravenous methotrexate with leucovorin rescue. The evidence favoring administration of intravenous methotrexate for children with ALL is critically reviewed. Despite its extensive use, high-dose intravenous methotrexate has not been proved conclusively to be more effective than less toxic, less labor intensive, and less costly methods of methotrexate administration.     

Attenuation of 2-methoxyethanol-induced testicular toxicity in the rat by simple physiological compounds

2-Methoxyethanol (2-ME) is an industrial solvent which is toxic to both male and female reproductive systems of laboratory animals. Earlier data have demonstrated that the developmental toxicity of 2-ME can be attenuated by simple physiological compounds such as serine, acetate, sarcosine, glycine, and D-glucose. The present experiments were designed to evaluate the same compounds for their ability to ameliorate the testicular toxicity that occurs in rats after 2-ME exposure. The extent of testicular damage was assessed by quantitating daily sperm production (DSP) on Day 24 following a single dose of 2-ME (6.6 mmol/kg, 500 mg/kg). Serine completely eliminated 2-ME-induced decreases in DSP, while glucose was without effect. Acetate, sarcosine, and glycine were of similar efficacy resulting in DSP that was significantly greater than that observed in rats which received 2-ME alone. Histopathological studies revealed that 2-ME treatment resulted in stage-specific degeneration of late stage pachytene spermatocytes 24 hr after treatment. No apparent degenerative changes occurred after concurrent treatment with serine. Similarly, serine also prevented the decreased number of spermatids in the lumina of the seminiferous tubules on Day 24 after 2-ME exposure alone. All of the compounds utilized in this study are linked to oxidation pathways involving tetrahydrofolic acid as a catalyst for one-carbon moiety transfer into purine and pyrimidine bases which are necessary precursors for DNA and RNA synthesis. The ability of these compounds to attenuate the testicular toxicity of 2-ME may result from their ability to donate one-carbon units which can be used in purine base biosynthesis. Reduced availability of bases would be expected to affect late stage pachytene spermatocytes which are known to be undergoing rapid RNA synthesis.     

Mechanism of leucovorin reversal of methotrexate cytotoxicity in human MCF-7 breast cancer cells

Previous studies have suggested that metabolic inhibition by methotrexate (MTX) is multifactorial and that cytotoxicity can be reversed by the reduced folate leucovorin. In this report we investigated the mechanism of leucovorin rescue in the MCF-7 human breast cancer cell line. Cells were exposed to various concentrations of MTX (0.5, 1.0, 3.0, and 10.0 microM) for 24 hr followed by rescue with labelled leucovorin (0.5 to 50 microM). The changes in the intracellular folate pools 24 hr following the addition of leucovorin were quantitated by high-pressure liquid chromatographic methods. The changes in the folate pools during rescue were compared with the ability of various concentrations of leucovorin to affect cellular rescue from MTX using a cloning assay. Our studies show that the total labelled intracellular folate pools increased in a log-linear fashion with respect to leucovorin exposure concentrations up to 100 microM. The degree of accumulation at a given leucovorin concentration was not significantly different in the absence or presence of MTX over the concentration range of 0.5 to 10 microM. Individual folate pool levels (tetrahydrofolate, 10-formyl tetrahydrofolate, 5-formyl tetrahydrofolate, 5-methyl tetrahydrofolate, and 5,10-methylene tetrahydrofolate) reached those present in cells not exposed to MTX at concentrations of leucovorin that were not adequate to rescue the MTX-treated cells. With exposure to concentrations of leucovorin capable of rescue, the individual folate pool levels were up to twelve times greater than those found in untreated cells, consistent with competition for catalytic activity at folate-dependent enzymes in addition to dihydrofolate reductase. The dihydrofolate pool also increased with increasing leucovorin concentration: but, unlike the reduced folates, this oxidized folate reached a maximal level that was dependent on the MTX concentration to which the cells had been exposed. This suggests that competition between MTX and leucovorin occurs at the level of dihydrofolate reductase via a competitive interaction with dihydrofolate in this intact cell system. The ability of leucovorin and its metabolites to compete with direct inhibitors of dihydrofolate reductase and other metabolically important folate-dependent enzymes appears to be associated with leucovorin rescue.     

Comparison of glycine metabolism in mouse lymphoma cells either sensitive or resistant to L-asparaginase

Previous work suggested a relationship between glycine metabolism and the effect of L-asparaginase upon tumor cells. Therefore, L5178Y (sensitive) or L5178Y/L-ASE (resistant) ascites lymphoma cells were incubated with 14C-labeled glyoxylate, glycine, serine, or asparagine, and the metabolism to other amino acids was measured by high performance liquid chromatography. Metabolic differences between the two cells lines were found. Under control conditions, the interconversion rate of glycine and serine via serine hydroxymethyltransferase (SHMT) was higher in sensitive than in resistant cells. The transformation rate of glyoxylate to serine was also higher in sensitive cells. These results may indicate a difference in the activity of SHMT. An alternate explanation would be that transport or diffusion of serine and glycine into sensitive cells is greater than into resistant cells. Several crucial metabolic differences were observed between the two cell types when L-asparaginase was added. A key difference is the decrease of glycine synthesis from glyoxylate observed in the sensitive cells compared to resistant cells which show no change. This suggests that asparagine is used for transamination of glyoxylate. Also, only sensitive cells appear to compensate for L-asparaginase-induced loss of glycine formation from glyoxylate by increasing glycine synthesis from serine. Alterations in sensitive tumor glycine metabolism may be an important function of L-asparaginase anticancer activity.     

Improved Pharmacy Department Workflow with New Method of Order Entry for Single-Agent,High-Dose Methotrexate

Purpose:

To determine whether a process change impacted the proportion of orders for single-agent, high-dose methotrexate entered by chemotherapy pharmacists instead of general pharmacy staff. Coordination of antiemetic premedication and leucovorin rescue with the new method of order entry was evaluated.

Methods:

Adults treated with single-agent, high-dose methotrexate were identified retrospectively. Order entry of methotrexate and ancillary medications was examined to determine whether the old or new method was used and whether it was performed by a chemotherapy pharmacist. The fundamental difference between the old and new methods for order entry is use of the “unscheduled” frequency of medication administration to replace the administration frequency of “once” with a specified date and time. Timing of antiemetic premedication and leucovorin rescue relative to methotrexate administration were tallied for the new method. Chi-square analysis was performed for the primary objective. Observational statistics were performed otherwise.

Results:

The number of evaluable encounters identified was 158. A chemotherapy pharmacist entered a greater proportion of orders when the new method was utilized (P < .0001). The proportion of orders entered by a chemotherapy pharmacist increased during the hours of 0700 and 2259 with the new method. Appropriate coordination of antiemetic and leucovorin administration was documented for 96% and 100% of cases with the new method of order entry.

Conclusion:

The proportion of orders for single-agent, high-dose methotrexate entered by a chemotherapy pharmacist was significantly greater with the use of the new method. Administration of antiemetic premedication and leucovorin rescue were appropriately coordinated with the use of the new method for order entry of single-agent, high-dose methotrexate.     

Intrathecal methotrexate neurotoxicity: clinical correlates and antidotal treatment

The neurotoxicity of methotrexate (MTX) is more severe when administered intrathecally (IT) than by the oral and intravenous (IV) routes, and has been reported even with a single administration of therapeutic doses of 12 or 15 mg. Prompt recognition and treatment are essential to improve the outcome after massive IT-MTX overdose. Treatment options include CSF drainage or CSF exchange, ventriculolumbar perfusion, IT corticosteroids to reduce CSF inflammation and IV leucovorin to reduce systemic toxicity. Toxicity resulting from IT injection of leucovorin is controversial. CSF drainage and exchange are particularly effective if performed soon after the overdose. In this paper we describe a protocol of treatment for severe cases of IT-MTX overdose in excess of 100 mg. The mainstay of treatment is dilution and removal from CSF of excessive methotrexate alongside with specific antidotal therapy.     

Acute cardiac toxicity associated with high-dose intravenous methotrexate therapy: case report and review of the literature

A 36-year-old woman was hospitalized for preoperative chemotherapy for osteosarcoma. She received intravenous fluids for 12 hours for volume expansion, then methotrexate 24 g (12 g/m2) over 6 hours. This was followed by intravenous leucovorin 200 mg over 1 hour. Two hours after the methotrexate infusion the patient developed chest pain and bradycardia. An electrocardiogram revealed sinus pauses, and telemetry recordings indicated a 4-beat run of ventricular tachycardia. A cardiac work-up consisting of cardiac enzyme level determination, two-dimensional echocardiography, and an adenosine technetium-99m tetrofosmin stress test was negative for structural and ischemic heart disease. The patient recovered without treatment and, approximately 2 weeks later, received a second course of methotrexate at half the dose without complication. One month later the patient received treatment with doxorubicin and cisplatin; 2 days later she died unexpectedly at home. Clinicians should be aware that high-dose methotrexate can cause cardiac symptoms and arrhythmias in previously healthy adults. This complication warrants attention and needs additional clinical investigation.     

Cytosolic factors that alter the metabolism of N,N-dimethyl-4-aminoazobenzene by rat liver microsomes

N,N-Dimethyl-4-aminoazobenzene (DAB), an azo dye carcinogen, is N-demethylated and 4'-hydroxylated by rat liver microsomes. Addition of hepatic cytosol to the microsomal system stimulated both pathways. This occurred in the presence of added NADPH or an NADPH-generating system. Cytosol was effective only when present prior to addition of substrate; no stimulation was seen when added after the reaction had begun. This suggested a direct effect on the microsomes rather than a chemical interaction with one or more metabolic intermediates of DAB. The degree of stimulation was somewhat different when using microsomes from phenobarbital- or beta-naphthoflavone-treated animals, implying a selectivity of the cytosolic effect for various isozymes of cytochrome P-450. Some loss of stimulatory activity occurred with dialysis. Activity was restored by adding back glutathione (GSH) which can stimulate DAB metabolism even in the absence of cytosol. DAB metabolism is also stimulated by EDTA. Although both EDTA and cytosol inhibit lipid peroxidation, cytosol stimulated DAB metabolism even in the presence of EDTA. Therefore, suppression of lipid peroxidation does not explain satisfactorily the cytosolic effect. Separation of cytosolic proteins by gel filtration revealed a factor which inhibits N-demethylation but not 4'-hydroxylation of DAB. Heating at 100 degrees partially inactivated the stimulatory activity. However, inhibitory activity was less susceptible to heat inactivation than was stimulatory activity. These results indicate that, in the whole cell, microsomal metabolism of xenobiotics is regulated to an appreciable extent by macromolecular cytosolic substances.     

Cardioprotective and chemosensitizing effects of novel Danshensu derivatives

OBJECTIVE To investigate the cardioprotective effect of novel danshensu derivatives against doxorubicin(Dox)cardiotoxicity and their synergistic anti-tumor effect with Dox on breast cancer cells.METHODS Two new Danshensu derivatives were synthesized by conjugation with tetramethylpyrizine and/or4-(3-thioxo-3 H-1,2-dithiol-4-yl)-benzoic acid and tested for protective effects against Dox induced cardiotoxicity in cell and zebrafish.H9c2 cardiomyoblasts were co-treated with Dox and Danshensu derivatives for 24 h and then were measured for cell viability and cytotoxicity by MTT and LDH assays.The expression levels of mitochondrial biogenesis related proteins PGC-1α,NRF-1and Nrf2 were detected by Western blotting and qPCR.Moreover,in a Dox-induced cardiotoxicity model of zebrafish,zebrafish embryos were treated with Dox for 36 h,followed by measurement of numerous ventricular function parameters including heart rate,stroke volume,cardiac output and fractional shortening.In addition,the synergistic anti-tumor effects of the Danshensu derivatives and Dox had been studied in MCF-7 breast cancer cells.The effects of the Danshensu derivatives on the cell death and metabolism of MCF-7 cells were measured using apoptosis assay and Seahorse Metabolic Analyzers respectively.RESULTS Our results showed that the Danshensu derivatives were more potent than the parental compounds in ameliorating Dox-induced cytotoxicity in H9c2 cells and significantly preserving stroke volume of heart function in Dox-treated zebrafish.Further mechanistic studies identified that the danshensu derivatives increased mitochondrial copy numbers and protein expressions of PGC-1α,NRF-1 and Nrf2 in H9c2 cells.In addition,the Danshensu derivatives enhanced Dox-induced apoptosis,and decreased glycolysis and mitochondrial function in MCF-7 tumor cells.CONCLUSION Our results revealed that two new Danshensu derivatives displayed promising cardioprotective effects against Dox induced cardiotoxicity both in vivo and in vitro,at least partially through activating mitochondrial biogenesis.Also,the new Danshensu derivatives potentiated the anticancer effects of Dox in breast tumor cells involving induction of glycolytic inhibition and mitochondrial dysfunction.     

Structural features of 4-amino antifolates required for substrate activity with mammalian folylpolyglutamate synthetase

The activity of a series of folic acid analogues as substrates for partially purified mouse liver folylpolyglutamate synthetase was determined and the effects of substituents on the binding to, and catalytic processes of, this enzyme were inferred. A 4-amino group improved substrate activity primarily by decreasing the apparent Km while N10-methyl substitution substantially diminished utilization as a substrate, again, by effects on Km. Isosteric replacement of N-10 altered substrate activity. A free alpha-carboxyl group in the amino acid side chain was required for catalysis as was the presence of the side chain amide carbonyl group. Modification of the amino acid side chain length profoundly affected activity. Several observations were made that may be relevant to chemotherapy with folate antimetabolites: 1) 7-hydroxymethotrexate was a substrate for this enzyme; 2) substrate activity and substrate inhibition were observed with CB 3717, a potent inhibitor of thymidylate synthase; 3) potent classical dihydrofolate reductase inhibitors were identified that were either not substrates for mouse liver folylpolyglutamate synthetase (e.g., 4-amino-4-deoxy-N10-methylpteroyl-L-alpha-aminoadipate) or were much better substrates than methotrexate for this enzyme (e.g., aminopterin); and 4) leucovorin and methotrexate appeared to be substrates for the same synthetase, but leucovorin saturated the reaction at much lower concentrations. These results have implications for the design of folylpolyglutamate synthetase inhibitors and for the selection of dihydrofolate reductase inhibitors that are either not polyglutamated or are efficiently polyglutamated in vivo.     

Endogenously formed norharman (beta-carboline) in platelet rich plasma obtained from porphyric rats

Porphyria was induced in adult male Wistar rats starved for 24 hr by SC injection of 400 mg/kg allylisopropylacetamide (AIA). The presence of porphyria was shown by measuring excretion of delta-aminolevulinic acid (delta-ALA) and porphobilinogen (PBG) into the urine during 24 hr after AIA administration. Plasma levels of glycine, serine and of a number of other amino acids were decreased in porphyric rats as compared to controls. Intraperitoneal injection of 2 mmol/kg serine 24 hr after AIA administration was used as an animal model for an acute psychosis, by measuring catalepsy scores 30 min after serine injection. The concentration of 5 different beta-carbolines in platelet rich plasma (PRP) was measured using an HPLC-fluorometric method. An increase in the concentration of norharman (NH) in PRP, ranging from 0.57 nmoles/l in control rats to 1.88 nmoles/l in serine treated porphyric rats was found. The catalepsy duration was exponentially correlated with the NH concentrations in PRP. It is concluded that an elevated conversion of serine into glycine via serine hydroxymethyltransferase (SHMT) may be responsible for the enhanced NH biosynthesis.     

Leflunomide: efficacy and safety in clinical trials for the treatment of rheumatoid arthritis

Leflunomide is a new disease modifying antirheumatic drug (DMARD) that inhibits lymphocyte proliferation by blocking dihydroorotate dehydrogenase (DHODH), the enzyme critical for the production of pyrimidine necessary for DNA synthesis. Through this mode of action, leflunomide inhibits the lymphocyte proliferation associated with the clonal expansion of T cells in rheumatoid arthritis (RA). In clinical trials, leflunomide was superior to placebo and comparable to sulfasalazine and methotrexate for improving both the signs and symptoms of RA. Leflunomide was also superior to placebo and sulfasalazine and comparable to methotrexate in overall improvement of physical function. Leflunomide was equivalent to methotrexate and sulfasalazine in retarding disease progression measured radiographically. Due to its unique mode of action in the treatment of RA, leflunomide shows value in combination therapy with methotrexate for patients refractory to methotrexate alone. The most common adverse reactions associated with leflunomide therapy include gastrointestinal symptoms, allergic reactions, reversible alopecia and elevated liver enzymes. Adverse events were generally mild to moderate, and resolved without complication. The results of phase II and phase III clinical trials indicate that leflunomide is a safe and efficacious drug for the treatment of RA.     

Sensitivity of zebrafish to environmental toxins implicated in Parkinson's disease

Parkinson's disease (PD) is characterized by the progressive loss of dopaminergic (DA) neurons in the substantia nigra and movement defects, including bradykinesia, tremor, and postural imbalance. Whereas the etiology and pathogenesis of PD is still poorly understood, studies in animal models are providing important insights. One valuable type of animal model for PD is established by treating animals with PD-inducing neurotoxins, including 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), rotenone, and paraquat. These neurotoxins are thought to inhibit mitochondrial complex I activity leading to oxidative stress, impaired energy metabolism, proteasomal dysfunction, and, eventually, dopamine neuronal loss. However, the genes and pathways that underlie the neurotoxicity of these agents are not known. In this study, we explored the effect of MPTP, rotenone, and paraquat in both adult and larval zebrafish, which are highly amenable to genetic analysis that can lead to the identification of the underlying genes and pathways. Here, we report that adult zebrafish display behavioral alterations, including decreased locomotor activity in response to MPTP, whereas larval zebrafish exhibited developmental, behavioral, and DA sensitivity to these agents. Taken together, these findings suggest that zebrafish could be a valuable model for genetically dissecting the molecular mechanisms underlying the neurotoxicity of PD-inducing agents.     

Brain ischemia induces serine phosphorylation of neuronal nitric oxide synthase by Ca^2＋ ／ calmodulin-dependent protein kinase II in rat hippocampus

INTRODUCTIONConstitutively expressed nitric oxide synthase(NOS) has three isoforms, including neuronal NOS(nNOS), inducible NOS (iNOS), and endothelial NOS(eNOS), which has been isolated and cloned[1]. NOScatalyzed L-arginine to generate L-citruline and nitricoxide (NO), a gaseous messenger molecule, the over-production of which plays a critical role in glutamate-induced neurotoxicity after ischemia[2]. In mutant micedeficient in nNOS and subsequent NO production, inf-arct s…