Areca nut extract suppresses T-cell activation and interferon-gamma production via the induction of oxidative stress.

Areca quid chewing is a major risk factor associated with oral submucous fibrosis (OSF) and oral cancer. Experimental evidence indicates that immune deterioration is associated with the pathophysiology of OSF and oral cancer. In addition, reactive oxygen species (ROS) is shown to play a role in the cytotoxic and genotoxic effect induced by areca nut extracts (ANE) in oral cells. The present studies investigated the effects of ANE on T-cell reactivity and the role of ROS in ANE effects. Treatment of splenocytes with ANE induced a marked cytotoxic effect, and suppressed the production of IL-2 and IFN-gamma, whereas the production of IL-4 was unaffected. The ANE-mediated cytotoxicity, and suppression of IFN-gamma and IL-2 production were attenuated by the presence of antioxidant N-acetyl-l-cysteine (NAC). Moreover, flow cytometric analysis demonstrated an increase in cellular ROS levels in splenic T-cells treated with ANE, which was also attenuated by the presence of NAC. Concordantly, the cellular level of glutathione was diminished by ANE in splenic T-cells pretreated with NAC. Collectively, these results demonstrated that ANE markedly suppressed T-cell activation and Th1 cytokine production, which was mediated, at least in part, by the induction of oxidative stress.

Biodistribution study of [99mTc] TRODAT-1 alone or combined with other dopaminergic drugs in mice with macroautoradiography.

A 99mTc labeled tropane derivative, [99mTc] TRODAT-1 (2beta-((N,N'-bis(2-mercaptoethyl) ethylene diamino)methyl), 3beta-(4-chlorophenyl) tropane), is a potential dopamine transporter (DAT) imaging agent for the central nervous system. To better understand the binding localization of [99mTc] TRODAT-1 both in the brain and the body, whole-body macroautoradiography (WBAR) was used in this study. The effect of DAT competing drugs, such as levadopa (L-DOPA), N-methyl-2beta-carbomethoxy-3beta-(4fluorophenyl)tropane (CFT, WIN 35,428) and methylphenidate, on the biodistribution of [99mTc] TRODAT-1 were also included in this study. Doses of 150 MBq [99mTc] TRODAT-1 were injected into normal male ICR mice through the caudal veins. For comparison, mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), L-DOPA, methylphenidate and CFT, respectively, were also investigated under the similar protocols. One and a half hours after [99mTc] TRODAT-1 injection, the mice were sacrificed. Whole-body autoradiography was performed immediately after sacrifice. Both frontal and sagittal sections showed that the liver and mucosa of stomach had the highest uptake of [99mTc] TRODAT-1. Other binding sites included the periphery of the spinal cord and the epithelium of the intestine. In the brain, autoradiographic imaging obtained from frontal sections showed symmetrical uptakes of [99mTc] TRODAT-1 in bilateral striata. Remaining binding sites include olfactory bulbs, thyroid gland, and salivary gland. The autoradiographic imaging obtained from sagittal sections showed a similar biodistribution. Mice treated with MPTP or L-DOPA showed no significant difference in the uptake of [99mTc] TRODAT-1 in bilateral striata, as compared to those of the control. In CFT or methylphenidate-treated mice, DAT binding sites were almost completely inhibited. These data showed that [99mTc] TRODAT-1 has potential clinical use for neurological investigation, such as Parkinson's and similar diseases.

The role of dopamine transporter imaging agent [99mTc]TRODAT-1 in hemi-parkinsonism rat brain.

This study aims to investigate the relationship between the determination of dopamine level by high performance liquid chromatography (HPLC) with electrochemical detection (ECD) and the detection of dopamine transporter (DAT) counts using autoradiography with DAT image agent [99mTc]TRODAT-1. For striatal lesions, pretreatment of 6-hydroxydopamine (6-OHDA) in the medial forebrain bundle shows that autoradiogaphic labeling of striatum region is reduced to near-background level. Using HPLC with ECD, unilateral 6-OHDA treatment is associated with significant (p < 0. 0002) reductions of dopamine levels. For the striatum of the 6-OHDA-lesioned side, dopamine content and DAT counts are reduced to 97% and 90%, respectively. Thus, our observation indicates a potential of using [99mTc]TRODAT-1 for the evaluation of animal DAT.

Evaluation of early-stage Parkinson's disease with 99mTc-TRODAT-1 imaging.

UNLABELLED: Parkinson's disease is a progressive neurodegenerative disorder characterized by a selective loss of dopamine in the striatum. Problems remain in the accurate diagnosis of Parkinson's disease. A 99mTc-labeled tropane derivative that binds to dopamine transporter with high selectivity is [2-[[2-[[[3-(4-chlorophenyl)-8-methyl-8-azabicyclo[3,2,1]oct-2-yl]methyl](2-mercaptoethyl)amino]ethyl]amino]ethanethiolato(3-)-N2,N2',S2,S2']oxo-[1R-(exo-exo)] (TRODAT-1). The purpose of this study was to investigate the potential usefulness of 99mTc-TRODAT-1 imaging in the evaluation of patients with early-stage Parkinson's disease. METHODS: Thirty-four patients with early-stage idiopathic Parkinson's disease were recruited. For all patients, the Parkinson's disease was stage 2 or less as assessed by the Hoehn and Yahr scale. Seventeen age-matched healthy volunteers (8 men, 9 women) served as controls. 99mTc-TRODAT-1 was prepared from a lyophilized kit. Brain SPECT imaging was performed between 165 and 195 min after injection, using a double-head camera equipped with fanbeam collimators. Specific uptake in the striatum and its subregions, including the putamen and caudate nucleus, was calculated and compared with that of the other sides and of healthy volunteers. RESULTS: A continuous reduction in specific striatal uptake of 99mTc-TRODAT-1 with increasing disease severity was found in Parkinson's disease patients (control vs. stage I vs. stage II, 1.98 vs. 1.62 vs. 1.22, respectively, P < 0.01). The changes were magnified by measurement of specific putaminal uptake (control vs. stage I vs. stage II, 1.81 vs. 1.27 vs. 0.94, respectively, P < 0.01). The mean values of specific putaminal uptake contralateral to the more affected limbs were significantly decreased compared with the ipsilateral sides in both stage I and stage II groups (1.02 vs. 1.49 for stage I and 0.73 vs. 1.14 for stage II, P < 0.01). Moreover, a significant loss of putaminal uptake ipsilateral to the symptoms was found in the stage I group compared with the healthy volunteers (1.49 vs. 1.81, P < 0.01). The difference became greater when the posterior putaminal uptakes were compared. No remarkable adverse reactions were found in either healthy volunteers or Parkinson's disease patients during or after imaging. CONCLUSION: For clinical practice, 99mTc-TRODAT-1 may serve as a useful imaging agent for the early detection of Parkinson's disease.

Development of a Tc-99m labeled sigma-2 receptor-specific ligand as a potential breast tumor imaging agent.

A novel in vivo imaging agent, 99mTc labeled [(N-[2-((3'-N'-propyl-[3,3,1]aza-bicyclononan-3alpha-yl)(2"-methoxy-5-methyl-phenylcarbamate)(2-mercaptoethyl)amino)acetyl]-2-aminoethanethiolato] technetium(V) oxide), [99mTc]2, displaying specific binding towards sigma-2 receptors was prepared and characterized. In vitro binding assays showed that the rhenium surrogate of [99mTc]2, Re-2, displayed excellent binding affinity and selectivity towards sigma-2 receptors (K(i) = 2,723 and 22 nM for sigma-1 and sigma-2 receptor, respectively). Preparation of [99mTc]2 was achieved by heating the S-protected starting material, 1, in the presence of acid, reducing agent (stannous glucoheptonate) and sodium [99mTc]pertechnetate. The lipophilic racemic mixture was successfully prepared in 10 to 50% yield and the radiochemical purity was >98%. Separation of the isomers, peak A and peak B, was successfully achieved by using a chiralpak AD column eluted with an isocratic solvent (n-hexane/isopropanol; 3:1; v/v). The peak A and peak B appear to co-elute with the isomers of the surrogate, Re-2, under the same HPLC condition. Biodistribution studies in tumor bearing mice (mouse mammary adenocarcinoma, cell line 66, which is known to over-express sigma-2 receptors) showed that the racemic [99mTc]2 localized in the tumor. Uptake in the tumor was 2.11, 1.30 and 1.11 %dose/gram at 1, 4 and 8 hr post iv injection, respectively, suggesting good uptake and retention in the tumor cells. The tumor uptake was significantly, but incompletely, blocked (about 25-30% blockage) by co-injection of "cold" (+)pentazocine or haloperidol (1 mg/Kg). A majority of the radioactivity localized in the tumor tissue was extractable (>60%), and the HPLC analysis showed that it is the original compound, racemic [99mTc]2 (>98% pure). The distribution of the purified peak A and peak B was determined in the same tumor bearing mice at 4 hr post iv injection. The tumor uptake was similar for both isomers, but the blood and peripheral tissue content for the isomer in peak B was higher than that for the isomer in peak A. It is evident that the isomer in peak A displayed significantly better tumor/blood and tumor/muscle ratios. The higher rate of in vivo metabolism was also confirmed by the higher thyroid uptake values for the isomer in peak B as compared to peak A. In summary, a 99mTc-labeled sigma receptor imaging agent, [99mTc]2, has demonstrated the feasibility of using a 99mTc-labeled agent for imaging sigma receptor expression in tumor cells. This is the first time a subtype-selective 99mTc-labeled agent for imaging sigma receptor sites is reported.

Differential diagnosis of Parkinson's disease and vascular parkinsonism by (99m)Tc-TRODAT-1.

UNLABELLED: The aim of this study was to use brain SPECT to differentiate vascular parkinsonism (VP) from Parkinson's disease. METHODS: Fourteen VP patients (age range, 59-87 y; mean age, 70 +/- 7.5 y), 30 Parkinson's disease patients (age range, 54-84 y; mean age, 65 +/- 8.8 y), and 26 healthy (control) individuals (age range, 50-85 y; mean age, 60 +/- 9 y) were examined. A 925-MBq (25 mCi) dose of (99m)Tc-TRODAT-1 was injected intravenously, and brain SPECT images were acquired 4 h after injection. The ratio of specific to nonspecific striatal (99m)Tc-TRODAT-1 binding was measured and compared. RESULTS: After a region-of-interest analysis of the images from VP patients, Parkinson's disease patients, and healthy volunteers was performed to obtain ratios of putamen to occipital and striatal to occipital binding as a measurement of specific binding to the dopamine transporters in these regions of the brain, where dopamine neurons are concentrated, the specific binding in the 14 VP patients was slightly lower than but not statistically different from that of the healthy individuals in both putamen and caudate areas. A significant decrease in uptake of (99m)Tc-TRODAT-1 in the striatum (P<0.01) was found in Parkinson's disease patients. Reduction of the uptake was more pronounced in the contralateral putamen of Parkinson's disease patients than that of VP patients (P<0.001). A significant bilateral striatal asymmetry was also observed in Parkinson's disease patients but not in VP patients (P< 0.01). CONCLUSION: Our findings clearly show that, for VP patients, (99m)Tc-TRODAT-1 SPECT is a reliable method to differentiate VP from Parkinson's disease. Further studies, including those to differentiate Parkinson's disease from arteriosclerotic parkinsonism and patients with both VP and Parkinson's disease, are needed to help rule out the possibility of Parkinson's disease as early as possible.

The optimal imaging time for [99Tcm]TRODAT-1/SPET in normal subjects and patients with Parkinson's disease.

Imaging of dopamine transporters (DATs) in the brain using [99Tcm]TRODAT-1 showed excellent pharmacokinetics for estimation of transporter concentrations. It has been reported that there may be differences in the binding kinetics of DAT radiotracers to DATs between normal subjects and patients with Parkinson's disease (PD). The aim of this study was to determine an optimal time point for (99Tcm]TRODAT-1 brain single photon emission tomography (SPET) acquisition that provides stable target to non-target ratios reflecting the DAT concentration in the brain. Serial [99Tcm]TRODAT-1 brain SPET images 2, 3 and 4 h after intravenous injection of [99Tcm]TRODAT-1 (925 MBq) were performed in five healthy subjects and nine PD patients. Regions of interests were drawn, and caudate/occipital (C/O) and putamen/occipital (P/O) specific to non-specific [99Tcm]TRODAT-1 binding ratios were calculated. The C/O and P/O ratios in healthy subjects showed consistent increases with time, but in PD patients, the C/O and P/O ratios of [99Tcm]TRODAT-1 reached a stable level at 3 h post-injection. There was a statistically significant difference (P < 0.001) between PD and normal subjects at 4 h post-injection for both the C/O and the P/O ratios. In conclusion, we recommend the acquisition of [99Tcm]TRODAT-1 SPET images at 4 h post-injection, as at this time point the C/O and P/O ratios can be used to discriminate between PD patients and healthy subjects.

Decreased dopamine transporter binding in Machado-Joseph disease.

UNLABELLED: The aim of this study was to use 99mTc-TRODAT-1 brain SPECT for investigation of the binding of dopamine transporter (DAT) in the nigrostriatal dopaminergic pathway of symptomatic Machado-Joseph disease (MJD) and to compare the results with the abnormal cytidylate, adenylate, and guanylate (CAG) expansion in the MJD1 gene and other clinical factors. METHODS: Ten symptomatic MJD patients (8 women, 2 men; age range, 20-71 y; mean age +/- SD, 36.4 +/- 10.6 y; mean duration of illness, 9.8 +/- 5.4 y) and 21 healthy volunteers (age range, 24-71 y; mean age, 47.6 +/- 20.1 y) were examined. Brain SPECT images were acquired 4 h after injection. The ratio of specific to nonspecific nigrostriatal 99mTc-TRODAT-1 binding was measured and compared with the clinical symptoms, duration of illness, and size of abnormal expanded CAG repeats. RESULTS: All nigrostriatal 99mTc-TRODAT-1 ratios were significantly lower in MJD patients than in healthy volunteers (P < 0.05). Discriminant function analysis of all MJD patients showed that the decreased binding of 99mTc-TRODAT-1 in the putamen was not significantly different from that in the caudate nucleus. Eight of 10 MJD patients had significantly decreased 99mTc-TRODAT-1 uptake. Of these 8, 2 had extrapyramidal signs and 6 had no obvious extrapyramidal signs. The other 2 patients, who had normal 99mTc-TRODAT-1 uptake, had no obvious extrapyramidal signs. CONCLUSION: Our findings indicate that 99mTc-TRODAT-1 brain SPECT is an appropriate method for evaluating damage to the nigrostriatal DAT in symptomatic MJD patients with and without extrapyramidal signs. The decreased binding of 99mTc-TRODAT-1 in the nigrostriatal dopaminergic pathway in symptomatic MJD patients correlates with the phenotype of extrapyramidal signs but not with the abnormal CAG repeat length, age at disease onset, or disease duration.

Novel gallium(III) complexes transported by MDR1 P-glycoprotein: potential PET imaging agents for probing P-glycoprotein-mediated transport activity in vivo.

BACKGROUND: Multidrug resistance (MDR) mediated by expression of MDR1 P-glycoprotein (Pgp) represents one of the best characterized barriers to chemotherapy in cancer patients. Positron emission tomography (PET) agents for analysis of Pgp-mediated drug transport activity in vivo would enable noninvasive assessment of chemotherapeutic regimens and MDR gene therapy. RESULTS: Candidate Schiff-base phenolic gallium(III) complexes were synthesized from their heptadentate precursors and gallium(III)acetylacetonate. Crystal structures demonstrated a hexacoordinated central gallium with overall trans-pseudo-octahedral geometry. Radiolabeled (67)Ga-complexes were obtained in high purity and screened in drug-sensitive (Pgp(-)) and MDR (Pgp(+)) tumor cells. Compared with control, lead compound 6. demonstrated antagonist-reversible 55-fold lower accumulation in Pgp-expressing MDR cells. Futhermore, compared with wild-type control, quantitative pharmacokinetic analysis showed markedly increased penetration and retention of 6. in brain and liver tissues of mdr1a/b((-/-)) gene disrupted mice, correctly mapping Pgp-mediated transport activity at the capillary blood-brain barrier and hepatocellular biliary cannalicular surface in vivo. CONCLUSIONS: These results indicate that gallium(III) complex 6. is recognized by MDR1 Pgp as an avid transport substrate, thereby providing a useful scaffold to generate (68)Ga radiopharmaceuticals for molecular imaging of Pgp transport activity in tumors and tissues in vivo using PET.

Pentavalent Tc-99m dimercaptosuccinic acid imaging of hepatocellular carcinoma.

Tc-99m (V)-dimercaptosuccinic acid (DMSA) has been used to image various kinds of tumors. However, from a review of the literature, it has never been applied to hepatocellular carcinoma (HCC). Low uptake of Tc-99m (V)-DMSA has been demonstrated in normal liver tissue, thus, Tc-99m (V)-DMSA may be useful for the detection of HCC. Nine male patients with focal nodular HCC were studied with sequential X-ray CT, Tc-99m (V)-DMSA and Tc-99m phytate liver scan. Our data showed that eight patients had increased uptake of Tc-99m (V) DMSA in HCC. Four cases demonstrated higher Tc-99m (V) DMSA uptake in HCC than in adjacent liver, and four cases demonstrated HCC uptake equal to liver uptake. One case showed no uptake of Tc-99m (V) DMSA in HCC. The detection sensitivity of Tc-99m (V)-DMSA was 88.9%. From our early results, Tc-99m (V)-DMSA is a readily available tumor imaging agent that appears to accumulate in HCC.

A trial study of leukocyte labeling with stabilized Tc-99m D,L-HMPAO by methylene blue and sodium phosphate buffer.

UNLABELLED: We attempted to label leukocytes with stabilized Tc-99m D,L-HMPAO by methylene blue and sodium phosphate buffer (S-HMPAO). METHODS: The results were compared with unstabilized Tc-99m D,L-HMPAO (U-HMPAO). U-HMPAO was obtained by reconstituting a commercial vial of D,L-HMPAO. Stabilization of the kit was performed by the addition of methylene blue. The leukocytes were labeled using a modified published method. The test samples of S-HMPAO and U-HMPAO were prepared immediately, and stood for 0.5, 1, 2, 4, and 6 h, respectively, at room temperature before analysis. RESULTS: In comparison with U-HMPAO: (1) the radiochemical purity of S-HMPAO was higher; (2) the labeling efficiencies of S-HMPAO labeled leukocytes were higher and consistent; (3) the viability of S-HMPAO labeled leukocytes was as high as the viability of U-HMPAO labeled leukocytes at any time; and (4) the percentages of disintegrated from S-HMPAO labeled leukocytes in plasma were lower. CONCLUSION: S-HMPAO is more stable than U-HMPAO and can provide higher leukocyte labeling efficiency. S-HMPAO, therefore, has the potential to replace U-HMPAO as a leukocyte-labeling agent.

The detection of nasopharynx carcinoma in technetium-99m (V) dimercaptosuccinic acid SPECT imaging.

Twenty-seven patients (24 men, 3 women; ages: 39-74 years) were diagnosed as having squamous cell carcinoma of the nasopharynx (NPC) as confirmed by pathologic findings of biopsies. In addition, three of the 27 patients had metastases to neck lymph nodes. The results of 2-4-hour SPECT images of the head and neck after intravenous injection of 15-20 mCi of Tc-99m (V) DMSA were compared with normal Tc-99m (V) DMSA images and CT of heads and necks. The results showed that of the 27 NPC cases, none of the patients had a significant uptake of Tc-99m (V) DMSA. However, in the three cases complicated with metastases of neck lymph nodes, the metastatic lesions could be detected by Tc-99m (V) DMSA. Our results challenge previous reports in which carcinomas of the head and neck were detected by Tc-99m (V) DMSA. The tumor-seeking agent Tc-99m (V) DMSA is not a good choice for the detection of NPC among malignancies of the head and neck.

Intravenous dipyridamole technetium-99m MMI myocardial perfusion scintigraphy for detection of coronary artery disease.

To evaluate the efficacy of stress technetium-99m MMI (N-2-methoxy-2-methyl propyl isonitrile) myocardial perfusion scintigraphy (Tc-99m MMI) using intravenous dipyridamole for detection of coronary artery disease (CAD), we collected 66 cases (53 men, 13 women, aged 40-79 years old) between Sept. 1990 and Oct. 1991. The cases were divided into two groups: group I involving 44 patients received coronary arteriography (CAG) examination without previous percutaneous transluminal coronary angioplasty (PTCA) or coronary artery bypass graft (CABG); group II embracing 22 patients received no CAG examination but all were suffering from old myocardial infarction (MI) evidenced by history and electrocardiography (ECG). All cases underwent Tc-99m MMI planar and single photon emission computed tomography (SPECT) both on intravenous dipyridamole stress and separate day rest tests. Of the 44 group I patients receiving both Tc-99m MMI and CAG, 35 (79.5%) were positive and 7 (16%) were negative by both tests and another 2 (4.5%) were positive by CAG only. Of those positive by both tests, 21 (60%) suffered from identical coronary arterial involvement, including 13 one-vessel disease, 5 double-vessel disease and 3 triple-vessel disease. Of group II patients, 14 suffered from old inferior wall (IW) MI, 6 from old anterior or anteroseptal wall (AW) MI, 1 from old lateral wall (LW) MI and another 1 from combined old anterior and lateral wall (ALW) MI by ECG. Of the 14 patients with IWMI by ECG, all suffered from right coronary artery (RCA) disease but 7 (50%) of them from multivessel disease (MVD) by Tc-99m MMI. 5 of the 6 patients with AWMI by ECG suffered from left anterior descending coronary artery (LAD) disease, but 3 of them from MVD by Tc-99m MMI. Both patients with LWMI and ALWMI by ECG suffered from triple-vessel disease by Tc-99m MMI. The sensitivity of Tc-99m MMI in detecting CAD in group I was 95%, the specificity was 100%. The sensitivity for detection of individual coronary artery disease in LAD, left circumflex coronary artery (LCX) and RCA was 96%, 45% and 89% in this order and the respective specificity was 94%, 100% and 88%. In group II the sensitivity was 95%. The overall sensitivity of intravenous dipyridamole Tc-99m MMI for detection of CAD in groups I and II was 95%; specificity was 100% and accuracy was 95%. In conclusion, stress Tc-99m MMI using intravenous dipyridamole is a valuable method for evaluation and detection of CAD.

Using technetium-99m (V) dimercaptosuccinic acid to detect malignancies from single solid masses in the lungs.

Fifty patients (43 male, 7 female, age 31-77 years) with single solid masses in their lungs based on the findings of a chest X-radiograph [40 malignancies: 5 small cell carcinoma (Ca), 17 epidermoid Ca, 12 adeno Ca, 6 undifferentiated large cell Ca] and 10 benign lesions underwent technetium-99m (V) dimercaptosuccinic acid [99m-(V)DMSA] scans to evaluate the usefulness of 99mTc-(V)DMSA in the detection of lung Ca with different cell types and benign lesions. Only 43% (17/40) of the malignancies in the lungs were detected by 99mTc-(V)DMSA, including 29% (5/17) epidermoid Ca, 50% (6/12) adeno Ca and 17% (1/6) undifferentiated large cell Ca of the lungs. However, all 5 cases of small cell Ca and 11 cases combined with bone metastasis were revealed by 99mTc-(V)DMSA. In addition, 3 of the 10 benign lesions (2 organizing pneumonias, 1 benign tumor) presented with an uptake of 99mTc-(V)DMSA. The diagnostic sensitivity, specificity and accuracy were 43%, 70% and 48%, respectively, in differentiating malignant from benign lesions for the single solid mass in the lungs. In conclusion, 99mTc-(V)DMSA is of little or no use in the differentiation of lung Ca from single solid masses in the lungs.