Tumor-induced osteomalacia: pre- and postoperative biochemical findings

A patient with late-onset hypophosphatemic osteomalacia was treated with oral supplements of phosphate (1.5 g/day) and calcitriol (1.5-3.0 micrograms/day) for 17 months, before a slowly growing tumor in the first metatarsal space became evident. Before treatment concentrations of inorganic phosphate (Pi) and calcitriol in serum and tubular reabsorption of phosphate (TRP) were very low, calcium and parathyroid hormone (PTH) in serum were normal, urinary cyclic adenosine monophosphate (cAMP) was strongly elevated. During the first weeks of conservative treatment urinary cAMP returned to normal; concomitantly there was a transient slight fall in PTH. Serum calcium was in the low normal range and did not significantly change during conservative therapy. During the further course PTH rose to pretreatment values, but urinary cAMP remained normal. When the dose of calcitriol was elevated to 3 x 1.0 micrograms/day, leading to slightly elevated serum concentrations of this substance, Pi in serum rose to the low normal range, but TRP remained low and bone pain, although improved, did not subside. The tumor was locally excised. Postoperatively calcitriol concentration became elevated within 48 hours and remained so for several weeks. The rise in calcitriol concentration preceded the elevation of Pi in serum, not, however, the increase of TRP. The elevation of urinary cyclic AMP before therapy may have been due to a direct action of the substance secreted by the tumor.     

Rejection prophylaxis with sequential OKT3 and CSA after kidney transplantation

Sixteen kidney transplant recipients received the IgG2a anti-CD3 monoclonal antibody OKT3 and azathioprine as rejection prophylaxis during the first two postoperative weeks. Concomitant immunosuppression consisted of low dose steroids while cyclosporine A therapy was instituted on day 12. Side effects included fever, bronchospasm, hypotension and diarrhoea. OKT3 caused T cell modulation resulting in CD3 dim +, CD4+ or CD8+, CD5+, WT31– and 11F2–cells. Anti-OKT3 antibodies were found in approximately 50% of the patients. The protocol induced a 100% patient and graft survival and a 81% actuarial freedom of rejection at 18 months. It prevented CsA associated nephrotoxicity in the direct postoperative phase.These beneficial effects outweighed the side effects of OKT3.     

Insulin Resistance and Dysregulation of Tryptophan–Kynurenine and Kynurenine–Nicotinamide Adenine Dinucleotide Metabolic Pathways

Insulin resistance (IR) underlines aging and aging-associated medical (diabetes, obesity, dyslipidemia, hypertension) and psychiatric (depression, cognitive decline) disorders. Molecular mechanisms of IR in genetically or metabolically predisposed individuals remain uncertain. Current review of the literature and our data presents the evidences that dysregulation of tryptophan (TRP)–kynurenine (KYN) and KYN–nicotinamide adenine dinucleotide (NAD) metabolic pathways is one of the mechanisms of IR. The first and rate-limiting step of TRP–KYN pathway is regulated by enzymes inducible by pro-inflammatory factors and/or stress hormones. The key enzymes of KYN–NAD pathway require pyridoxal-5-phosphate (P5P), an active form of vitamin B₆, as a cofactor. Deficiency of P5P diverts KYN–NAD metabolism from production of NAD to the excessive formation of xanthurenic acid (XA). Human and experimental studies suggested that XA and some other KYN metabolites might impair production, release, and biological activity of insulin. We propose that one of the mechanisms of IR is inflammation- and/or stress-induced upregulation of TRP–KYN metabolism in combination with P5P deficiency-induced diversion of KYN–NAD metabolism towards formation of XA and other KYN derivatives affecting insulin activity. Monitoring of KYN/P5P status and formation of XA might help to identify subjects at risk for IR. Pharmacological regulation of the TRP–KYN and KYN–NAD pathways and maintaining of adequate vitamin B₆ status might contribute to prevention and treatment of IR in conditions associated with inflammation/stress-induced excessive production of KYN and deficiency of vitamin B₆, e.g., type 2 diabetes, obesity, cardiovascular diseases, aging, menopause, pregnancy, and hepatitis C virus infection.     

The Role of Tryptophan Catabolism along the Kynurenine Pathway in Acute Ischemic Stroke

Post-stroke inflammation may induce upregulation of the kynurenine (KYN) pathway for tryptophan (TRP) oxidation, resulting in neuroprotective (kynurenic acid, KA) and neurotoxic metabolites (3-hydroxyanthranillic acid, 3-HAA). We investigated whether activity of the kynurenine pathway in acute ischemic stroke is related to initial stroke severity, long-term stroke outcome and the ischemia-induced inflammatory response. Plasma concentrations of TRP and its metabolites were measured in 149 stroke patients at admission, at 24 h, at 72 h and at day 7 after stroke onset. We evaluated the relation between the KYN/TRP ratio, the KA/3-HAA ratio and stroke severity, outcome and inflammatory parameters (C-reactive protein (CRP), erythrocyte sedimentation rate (ESR) and neutrophil/lymphocyte ratio (NLR)). KYN/TRP but not KA/3-HAA correlated with the NIHSS score and with the infarct volume. Patients with poor outcome had higher mean KYN/TRP ratios than patients with more favourable outcome. The KYN/TRP ratio at admission correlated with CRP levels, ESR and NLR. The activity of the kynurenine pathway for tryptophan degradation in acute ischemic stroke correlates with stroke severity and long-term stroke outcome. Tryptophan oxidation is related to the stroke-induced inflammatory response.     

Changes of certain parameters of renal function in patients with allogenic bone marrow transplantation]

Serum creatinine and urea changes as well as protein, erythrocytes, and leucocytes excretion with the urine in allogenic bone marrow recipients are discussed. An increase in serum creatinine and urea together with proteinuria and hematuria were noted in 1/3 of patients treated with busulphan and cyclophosphamide prior to bone marrow transplantation. In cases of bone marrow graft rejection or post-transplantation complications no abnormalities in the value of investigated parameters were noted, especially immediately before death. Possible causes of such renal function disorders are also discussed.     

Metabolism of [5-3H]Kynurenine in the Rat Brain In Vivo: Evidence for the Existence of a Functional Kynurenine Pathway

Abstract: The incorporation of tritium label into quinolinic acid (QUIN), kynurenic acid (KYNA), and other kynurenine (KYN) pathway metabolites was studied in normal and QUIN-lesioned rat striata after a focal injection of [5-³H]KYN in vivo. The time course of metabolite accumulation was examined 15 min to 4 h after injection of [5-³H]KYN, and the concentration dependence of KYN metabolism was studied in rats killed 2 h after injection of 1.5–1,500 µ M [5-³H]KYN. Labeled QUIN, KYNA, 3-hydroxykynurenine (3-HK), 3-hydroxyanthranilic acid, and xanthurenic acid (XA) were recovered from the striatum in every experiment. Following injection of 15 µ M [5-³H]KYN, a lesion-induced increase in KYN metabolism was noted. Thus, the proportional recoveries of [³H]KYNA (5.0 vs. 1.8%), [³H]3-HK (20.9 vs. 4.5%), [³H]XA (1.5 vs. 0.4%), and [³H]QUIN (3.6 vs. 0.6%) were markedly elevated in the lesioned striatum. Increases in KYN metabolism in lesioned tissue were evident at all time points and KYN concentrations used. Lesion-induced increases of the activities of kynurenine-3-hydroxylase (3.6-fold), kynureninase (7.6-fold), kynurenine aminotransferase (1.8-fold), and 3-hydroxyanthranilic acid oxygenase (4.2-fold) likely contributed to the enhanced flux through the pathway in the lesioned striatum. These data provide evidence for the existence of a functional KYN pathway in the normal rat brain and for a substantial increase in flux after neuronal ablation. This method should be of value for in vivo studies of cerebral KYN pathway function and dysfunction.     

Pituitary-adrenocortical axis, serum serotonin and biochemical response after halothane or isoflurane anaesthesia in rabbits

To document the changes in serum serotonin, adrenocorticotrophic hormone (ACTH), corticosterone levels and select biochemical parameters in response to inhalant anaesthesia, 20 New Zealand White (NZW) rabbits were assigned to two treatment groups: halothane and isoflurane. Induction of anaesthesia was achieved using a face mask (3.5% halothane and 4.5% isoflurane in oxygen) followed by endotracheal intubation and maintenance of anaesthesia for 30 min (1.5% halothane and 2.5% isoflurane in oxygen). Blood samples were obtained before anaesthetic induction, and at 1, 10, 30, 60, 120 min and 24, 48 and 72 h after endotracheal intubation. Serum serotonin and corticosterone levels were measured by competitive enzyme immunoassay, ACTH by radioimmunoassay. Serum glucose, alanine aminotransferase (ALT), aspartate aminotransferase (AST), alkaline phosphatase (ALP), blood urea nitrogen (BUN) and creatinine levels were measured using an automated analyser. Significant increases in serum ACTH and corticosterone levels occurred after halothane administration while serum serotonin levels did not change. An increase in serum corticosterone and serotonin levels occurred in the isoflurane group but no changes in ACTH concentrations were detected. Administration of halothane significantly increased serum glucose, ALT, AST, BUN and creatinine levels. After isoflurane administration, there was a significant increase in serum glucose, AST, BUN and creatinine levels. Based on these results, halothane stimulates the hypothalamic-pituitary-adrenal axis to a greater extent than isoflurane, but isoflurane increases serum serotonin levels. Both anaesthetic agents alter select biochemical parameters. These results should be taken into account when blood samples are evaluated in treated isoflurane or halothane anaesthetized rabbits.     

Measurement of serum amyloid A protein concentrations as test of renal allograft rejection in patients with initially non-functioning grafts.

Serum amyloid A protein concentrations were monitored in 10 renal transplant recipients who required dialysis after transplantation because of an initially non-functioning graft. Fifteen rejection episodes were identified by repeated fine needle aspiration biopsies of the grafts. All rejections were characterised by pronounced increases in serum amyloid A concentrations, the mean peak value being 363 (SE 57) mg/1 as compared with a mean preoperative concentration of 14 (5) mg/1. The rise in concentrations preceded the start of anti-rejection treatment by an average of 2.5 days in eight of the rejection episodes, in five episodes it occurred the same day, and in two episodes it occurred the next day. With exclusion of the predictable surgery induced rise in values, which peaked on the second postoperative day, there were 17 increases in amyloid A concentrations peaking at greater than or equal to 100 mg/1; in two cases they were not related to documented rejection. These findings show that measurements of serum amyloid A concentration provide a valuable non-invasive aid in identifying acute renal allograft rejection, including that in patients whose graft does not function initially.     

Determination of serum kynurenine and hepatic tryptophan dioxygenase activity by high-performance liquid chromatography

The status of the oxidative metabolism of L-tryptophan is usually evaluated by the determination of tryptophan metabolites in serum or urine and/or the activities of various oxidative enzymes in tissues. I have developed assays for serum kynurenine and hepatic tryptophan dioxygenase (TDO) activity based on the determination of kynurenine (KYN) by isocratic, reverse phase HPLC with spectrophotometric detection at 365 nm. Sample pretreatment prior to HPLC requires little more than perchloric acid precipitation of serum or a TDO incubation mixture. The analytical recovery for the serum assay was 101 +/- 2%, while the run-to-run coefficient of variation at normal KYN levels was approximately 8%. Serum KYN levels in 40 apparently healthy fasting humans were normally distributed and ranged from 0.27 to 0.69 microgram/ml (mean +/- SD: 0.47 +/- 0.1). Serum KYN in predialysis specimens from a group of 20 patients with chronic renal failure demonstrated a highly significant increase (mean +/- SD: 0.83 +/- 0.35 microgram/ml; P less than 0.001) as compared to the reference population. It is possible that such an increase might contribute to the pathophysiology of the uremic state. The analytical recovery of KYN from TDO incubation mixtures was approximately 90%. There was no evidence for the onward metabolism of KYN during the assay of whole liver homogenates. The mean (+/- SD) TDO activity of rat liver homogenates preincubated with ascorbate and hematin was 2.3 +/- 0.8 mumol/h/g wet wt (30 degrees C). The sensitivity, specificity, and convenience of these two methods suggest that they are suitable for routine use in the investigation of the biology and pathology of oxidative tryptophan metabolism.     

Use of EQUORAL in de novo renal transplant recipients

This paper presents our experience to date with using a cyclosporine formulation Equoral (IVAX Pharmaceuticals) together with mycophenolate mofetil plus a steroid immunosuppressive regimen in the treatment of de novo renal transplant recipients. Ten cadaveric donor renal transplant recipients of mean age 51.6 years (range 37-66) were followed up over 6 months for the development of rejection attacks and side effects. All patients received prednisolone, mycophenolate mofetil (1 g/day during the first 5 days posttransplant and then 20 mg/kg/day) plus cyclosporine (3 mg/kg/day). Biopsy proven acute rejection episodes were observed in 2 out of 10 patients (20%). Six months patient as well as renal graft survival rate was 100%. The development of graft function was immediate after transplantation. The mean serum creatinine levels were gradually decreased. Over the 6-month posttransplant period, the function of the graft was satisfactory and stable. The majority of observed adverse events were those commonly reported with the use of cyclosporine and they resolved with a reduction in cyclosporine dose. Equoral treatment demonstrated an acceptable safety profile with maintenance of adequate renal function without incidence of malignancy/lymphoproliferative disease or serious infections. In conclusion, Equoral plus mycophenolate mofetil immunosuppression seems effective and safe on terms acute rejection rates, patient and renal graft survival rates and side profiles.     

Involvement of the Kynurenine Pathway in Human Glioma Pathophysiology

The kynurenine pathway (KP) is the principal route of L-tryptophan (TRP) catabolism leading to the production of kynurenine (KYN), the neuroprotectants, kynurenic acid (KYNA) and picolinic acid (PIC), the excitotoxin, quinolinic acid (QUIN) and the essential pyridine nucleotide, nicotinamide adenine dinucleotide (NAD⁺). The enzymes indoleamine 2,3-dioxygenase-1 (IDO-1), indoleamine 2,3-dioxygenase-2 (IDO-2) and tryptophan 2,3-dioxygenase (TDO-2) initiate the first step of the KP. IDO-1 and TDO-2 induction in tumors are crucial mechanisms implicated to play pivotal roles in suppressing anti-tumor immunity. Here, we report the first comprehensive characterisation of the KP in 1) cultured human glioma cells and 2) plasma from patients with glioblastoma (GBM). Our data revealed that interferon-gamma (IFN-γ) stimulation significantly potentiated the expression of the KP enzymes, IDO-1 IDO-2, kynureninase (KYNU), kynurenine hydroxylase (KMO) and significantly down-regulated 2-amino-3-carboxymuconate semialdehyde decarboxylase (ACMSD) and kynurenine aminotransferase-I (KAT-I) expression in cultured human glioma cells. This significantly increased KP activity but significantly lowered the KYNA/KYN neuroprotective ratio in human cultured glioma cells. KP activation (KYN/TRP) was significantly higher, whereas the concentrations of the neuroreactive KP metabolites TRP, KYNA, QUIN and PIC and the KYNA/KYN ratio were significantly lower in GBM patient plasma (n = 18) compared to controls. These results provide further evidence for the involvement of the KP in glioma pathophysiology and highlight a potential role of KP products as novel and highly attractive therapeutic targets to evaluate for the treatment of brain tumors, aimed at restoring anti-tumor immunity and reducing the capacity for malignant cells to produce NAD⁺, which is necessary for energy production and DNA repair.     

Th1/Th2 cytokines and ICAM-1 levels post-liver transplant do not predict early rejection

Th1 derived cytokines IFN-gamma and IL-2, Th2 cytokine IL-4, and ICAM-1 have been implicated in liver allograft rejection. In order to determine whether monitoring of cytokine profiles during the first days post-liver transplant can predict early rejection we measured IFN-gg, IL-2, sIL-2 receptor, IL-4 and ICAM-1 in 22 patients, in plasma samples obtained within 4 h after liver perfusion (baseline) and between postoperative days (POD) 3-6. ICAM-1 and sIL-2R levels at POD 3-6 were significantly higher than at baseline but did not differ in presence or absence of rejection. Mean percentage increase of ICAM-1 levels was significantly lower in patients with Muromonab-C3 Orthoclone OKT3 (J.C. Health Care) (OKT3) whereas percentage increase of sIL-2R levels was higher in OKT3-treated patients. IFN-gamma levels at POD 3-6 increased from baseline while IL-4 levels were unchanged. Levels of IFN-gamma, IL-4 and their ratios did not correlate with rejection or immunosuppressive therapy. Thus, Th1/Th2 cytokine monitoring during the first week post-transplant does not predict early rejection and immunosuppressive therapy is the predominant factor affecting ICAM and sIL-2R levels after liver transplantation.     

Influence of gender on tamoxifen-induced biochemical changes in serum of rats

Tamoxifen, the widely prescribed drug in the prevention and therapy of breast cancer, may cause side effects which may be influenced by gender. The present study was undertaken to investigate the impact of gender on tamoxifen-induced toxic and biochemical changes following oral administration of tamoxifen at high dose level of 20 mg/kg once daily for a 2-week period in both male and female rats. The results showed marked increases in serum activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in female rats. In contrast, treatment with tamoxifen in male animals significantly decreased the activity of ALT, with a tendency for a decrease in serum AST levels. In female rats, a significant reduction in the serum activity of acid phosphatase (ACP) was noted, compared with a non-significant decrease in males. Non-significant changes in serum levels of alkaline phosphatase (ALP) were seen in both sexes. Tamoxifen lowered serum contents of total lipid and total cholesterol in both male and female rats. Serum levels of triglycerides were reduced in female rats as compared to a non-significant decrease in male animals. The serum albumin concentration was decreased in both male and female rats, while total protein was decreased only in female animals. Tamoxifen markedly increased serum levels of creatinine in female rats, compared with a non-significant rise in males. Total serum contents of calcium were similarly reduced in both males and females. This is the first study which points to gender-related differences in tamoxifen-induced toxic and metabolic changes in rats. The results indicated that females are more susceptible than males to tamoxifen toxicity, probably due to the ability of tamoxifen to antagonize the action of estrogen in females.     

Optimization of Zn2+-containing mobile phase for simultaneous determination of kynurenine,kynurenic acid and tryptophan in human plasma by high performance liquid chromatography

In the present work we have developed a standard-addition HPLC method using a mobile phase containing low concentration of ZnAc₂ to determine physiological level of kynurenine (KYN), kynurenic acid (KYNA) and tryptophan (TRP) in human plasma simultaneously. The method greatly improved the sensitivity of KYNA, the resolution of KYNA and TRP, and avoided clotting risk caused by high concentration of ZnAc₂ in mobile phase. Samples were deproteinized by addition of equal volume of 0.6 mol/L HClO₄. Analytes in supernatants were separated by an Agilent HC-C18 (2) analytical column; an aqueous mobile phase containing 20 mmol/L NaAc, 3 mmol/L ZnAc₂ and 7% acetonitrile at flow rate of 1.0 mL/min. Detections were performed by a variable wavelength detector at wavelength 365 nm for KYN and a fluorescence detector at wavelengths excitation 344 nm and emission 398 nm for KYNA and TRP. Good linear responses were found with r² > 0.999 for all analytes within the concentration range of physiological levels. The limit of detection of the developed method was 0.03 μmol/L, 0.9 nmol/L and 0.4 μmol/L for KYN, KYNA and TRP respectively. Recoveries from spiked human plasma were 95.4–99.7% for KYN, 98.9–104% for KYNA and 96.5–100.2% for TRP. All CVs for the repeatability and intermediate precision were less than 5%. We conclude that the developed method is helpful for the research investigations in KYN pathway of TRP metabolism.     

OKT4/8 ratio in the blood and in the graft during episodes of human renal allograft rejection

We have analyzed the frequency of T helper (Th) and T suppressor/killer (Ts/k) lymphocytes in the blood and in the renal allograft during episodes of rejection and during quiescence. Monoclonal OKT4 and OKT8 antibodies were used to mark the Th and Ts/k cells, respectively. Density centrifugation-separated mononuclear leukocytes and FACS IV cell sorter or the Staphylococcus aureus rosette assay were used to determine the ratio in the blood, with concordant results. Fine needle aspiration biopsy (FNAB) and the Staph. assay were used to demonstrate the lymphocyte subtypes in the graft. The mean OKT4/8 ratio in the blood was significantly lower in the transplant recipients than in healthy controls (1.1 +/- 0.7 vs 1.8 +/- 0.2, respectively, P = 0.000). The individual variation was, however, high and no correlation between the OKT4/8 ratio in the blood and the inflammatory episodes in situ was observed. During 19 of the 25 episodes of inflammation, the dominant lymphocyte subtype in the graft was the Ts/k cell. In the remaining six cases it was the Th cell. All rejection episodes of the former type were reversible, in the latter type, four out of six were irreversible.     

CISPLATIN-INDUCED VOMITING DEPENDS ON CIRCADIAN TIMING

We examined whether the clock time of cisplatin plus antiemetic and diuretic administration affects the amount of cisplatin-associated emesis and severity of renal toxicity. We treated 22 patients with urogenital cancer with two courses of chemotherapy containing 70 mg/m² of cisplatin. Cisplatin together with furosemide was administered in the morning (05:00) or evening (17:00) during two courses 1 month apart in a crossover fashion. Ondansetron was given either before or after cisplatin to control nausea and vomiting. The number of vomiting episodes, serum creatinine, serum urea nitrogen (BUN), creatinine clearance, and urinary β-N-acetyl glucosamidase (NAG) concentration were evaluated before and after each treatment course. Regardless of the timing of ondansetron, morning compared to evening cisplatin was always associated with greater vomiting in the first treatment course. However, prophylactic administration of ondansetron markedly diminished the impact of the clock time of cisplatin administration. Serum creatinine transiently decreased rather than increased 14 days after cisplatin and furosemide administration, while NAG excretion increased 3 days after cisplatin and furosemide administration. In the first course, serum creatinine levels were similar regardless of the clock time of cisplatin and furosemide administration. However, in the second course, serum creatinine rose in patients given evening cisplatin and furosemide, while it remained unchanged in those given morning cisplatin and furosemide. Moreover, the first course morning cisplatin and furosemide treatment was associated with less change in NAG excretion (less kidney toxicity) than the first course of evening cisplatin and furosemide treatment. The second course evening cisplatin and furosemide treatment was associated with an increase in NAG excretion compared to the first course of treatment, while morning cisplatin and furosemide treatment in the second course showed less change in NAG excretion compared to the first course. The clock time of cisplatin administration had an impact on the frequency of emesis. Prophylactic ondansetron, however, diminished the time-of-day dependency of cisplatin-induced vomiting. Administration of cisplatin and furosemide in the morning rather than evening appears to cause less renal damage, and this damage may be further reduced with aggressive hydration and routine administration of furosemide. (Chronobiology International, 18(5), 851-863, 2001)     

CISPLATIN-INDUCED VOMITING DEPENDS ON CIRCADIAN TIMING

We examined whether the clock time of cisplatin plus antiemetic and diuretic administration affects the amount of cisplatin-associated emesis and severity of renal toxicity. We treated 22 patients with urogenital cancer with two courses of chemotherapy containing 70 mg/m² of cisplatin. Cisplatin together with furosemide was administered in the morning (05:00) or evening (17:00) during two courses 1 month apart in a crossover fashion. Ondansetron was given either before or after cisplatin to control nausea and vomiting. The number of vomiting episodes, serum creatinine, serum urea nitrogen (BUN), creatinine clearance, and urinary β-N-acetyl glucosamidase (NAG) concentration were evaluated before and after each treatment course. Regardless of the timing of ondansetron, morning compared to evening cisplatin was always associated with greater vomiting in the first treatment course. However, prophylactic administration of ondansetron markedly diminished the impact of the clock time of cisplatin administration. Serum creatinine transiently decreased rather than increased 14 days after cisplatin and furosemide administration, while NAG excretion increased 3 days after cisplatin and furosemide administration. In the first course, serum creatinine levels were similar regardless of the clock time of cisplatin and furosemide administration. However, in the second course, serum creatinine rose in patients given evening cisplatin and furosemide, while it remained unchanged in those given morning cisplatin and furosemide. Moreover, the first course morning cisplatin and furosemide treatment was associated with less change in NAG excretion (less kidney toxicity) than the first course of evening cisplatin and furosemide treatment. The second course evening cisplatin and furosemide treatment was associated with an increase in NAG excretion compared to the first course of treatment, while morning cisplatin and furosemide treatment in the second course showed less change in NAG excretion compared to the first course. The clock time of cisplatin administration had an impact on the frequency of emesis. Prophylactic ondansetron, however, diminished the time-of-day dependency of cisplatin-induced vomiting. Administration of cisplatin and furosemide in the morning rather than evening appears to cause less renal damage, and this damage may be further reduced with aggressive hydration and routine administration of furosemide. (Chronobiology International, 18(5), 851–863, 2001)     

Effect of kynurenine loading on quinolinic acid production in the rat: studies in vitro and in vivo

In vitro and in vivo techniques were used to examine the production and subsequent fate of the endogenous excitotoxin quinolinic acid (QUIN) following administration of its bioprecursor L-kynurenine (KYN). Incubation of liver slices in the presence of 10-1000 microM KYN resulted in a dose- and time-dependent release of QUIN into the incubation medium. Less than 15% of total QUIN produced was recovered from the tissue. In vivo experiments, performed with a microdialysis probe inserted in the jugular vein of anesthetized rats, showed that injection of KYN (20-600 mg/kg, i.v.) causes rapid and dose-dependent increases in the serum level of QUIN. Peak QUIN concentrations in serum dialysates were reached 75 minutes following KYN administration. Longer lasting increases were detected following the administration of pyrazinamide (20 mg/kg, i.p.), an indirectly acting stimulator of QUIN biosynthesis in the periphery. The data demonstrate the feasibility of assessing the mechanisms of QUIN production and disposition in experimental paradigms which can be expected to allow insights into the function and possible dysfunction of QUIN in the brain.     

Metabonomics study of the acute graft rejection in rat renal transplantation using reversed-phase liquid chromatography and hydrophilic interaction chromatography coupled with mass spectrometry

Acute graft rejection is one of the most common and serious post complications in renal transplantation, noninvasive diagnosis of acute graft rejection is essential for reducing risk of surgery and timely treatment. In this study, a non-targeted metabonomics approach based on ultra performance liquid chromatography (UPLC) coupled with quadrupole time-of-flight mass spectrometry (MS) is used to investigate the effect of acute graft rejection in rat renal transplantation on metabolism. To collect more metabolite information both hydrophilic interaction chromatography and reversed-phase liquid chromatography were used. Using the partial least squares-discriminant analysis, we found that the change of metabonome in a sham-operated group and a non-graft rejection group had a similar trend, while that of the acute graft rejection group was clearly different. Several discriminating metabolites of the acute graft rejection were identified, including creatinine, phosphatidyl-cholines, lyso-phosphatidylcholines, carnitine C16:0, free fatty acids and indoxyl sulfate etc. These discriminating metabolites suggested that acute graft rejection in renal transplantation can lead to the accumulation of creatinine in the body, and also the abnormal metabolism of phospholipids. These findings are useful to understand the mechanisms of the rejection, it also means that a UPLC-MS metabonomic approach is a suitable tool to investigate the metabolic abnormality in the acute graft rejection in renal transplantation.     

Biogenic monoamine levels in the central nervous system of the sea hare,Aplysia kurodai

1. By using a three-dimensional-coulometric HPLC system, biogenic monoamines and their metabolites were quantified simultaneously in the central nervous system of the sea hare, Aplysia kurodai.2. Precursor amino acids, tyrosine-4 (TYR-4) and tryptophan (TRP), and dopamine (DA), 3, 4-dihydroxyphenylacetic acid (DOPAC) and 5-hydroxytryptamine (5-HT) were detected in all the ganglia examined.3. Levels of these compounds in the cerebral, pedal and parieto-visceral ganglia were higher than those of the other ganglia examined.4. In some ganglia, epinephrine (E), 3-O-methyldopa (30MD), 3-methoxytyramine (3-MT), dihydroxyphenylethleneglycol (DOPEG), metanephrine (MN), vanillic acid (VA), octopamine (OCT), kynurenine (KYN) and 5-hydroxyindoleacetic acid (5-HIAA) were also detected.5. The main metabolic pathways of biogenic monoamines were shown to be TYR-4DADOPAC and TRP5-HT5-HIAA. Furthermore, following five pathways were also suggested to be present; TYR-4DAEMNVA, TYR-4TYRAOCT, TYR-43OMD, DA3-MT. EDOPEG and TRPKYN.