Anti-metastasis activity of 5,4'-dihydroxy 6,8-dimethoxy 7-O-rhamnosyl flavone from Indigofera aspalathoides Vahl on breast cancer cells.

Breast cancer presents a significant challenge due to its high rates of illness and mortality, necessitating more effective treatment approaches. While traditional treatments offer some benefits, they often lack precision in targeting cancer cells and can inadvertently harm healthy tissues. This study aims to investigate the cytotoxic effects and molecular mechanism of 5,4'-dihydroxy-6,8-dimethoxy-7-O-rhamnosyl flavone (DDR), extracted from Indigofera aspalathoides Vahl, on breast cancer cells (MDA-MB-231). Through various in vitro assays including wound healing, invasion, Western blotting, and immunofluorescence, the impact of DDR on epithelial-mesenchymal transition (EMT) and metastasis was evaluated. Treatment of MDA-MB-231 cells with different DDR concentrations (0-10 µg/mL) resulted in a significant decrease in invasion and migration, accompanied by the downregulation of metastasis-related proteins including VEGF, uPAR, uPA, and MMP-9. DDR treatment also hindered EMT by upregulating E-cadherin and downregulating N-cadherin, Slug, Twist, and Vimentin. Additionally, inhibition of the PI3K/AKT signaling pathway and downregulation of the NF-кB pathway were observed. These findings highlight the potential of DDR as a valuable source of natural compounds with promising anticancer properties, offering opportunities for the development of novel cancer therapies.

Geraniol attenuates oxidative stress and neuroinflammation-mediated cognitive impairment in D galactose-induced mouse aging model.

D-galactose (D-gal) administration was proven to induce cognitive impairment and aging in rodents' models. Geraniol (GNL) belongs to the acyclic isoprenoid monoterpenes. GNL reduces inflammation by changing important signaling pathways and cytokines, and thus it is plausible to be used as a medicine for treating disorders linked to inflammation. Herein, we examined the therapeutic effects of GNL on D-gal-induced oxidative stress and neuroinflammation-mediated memory loss in mice. The study was conducted using six groups of mice (6 mice per group). The first group received normal saline, then D-gal (150 mg/wt) dissolved in normal saline solution (0.9%, w/v) was given orally for 9 weeks to the second group. In the III group, from the second week until the 10th week, mice were treated orally (without anesthesia) with D-gal (150 mg/kg body wt) and GNL weekly twice (40 mg/kg body wt) four hours later. Mice in Group IV were treated with GNL from the second week up until the end of the experiment. For comparison of young versus elderly mice, 4 month old (Group V) and 16-month-old (Group VI) control mice were used. We evaluated the changes in antioxidant levels, PI3K/Akt levels, and Nrf2 levels. We also examined how D-gal and GNL treated pathological aging changes. Administration of GNL induced a significant increase in spatial learning and memory with spontaneously altered behavior. Enhancing anti-oxidant and anti-inflammatory effects and activating PI3K/Akt were the mechanisms that mediated this effect. Further, GNL treatment upregulated Nrf2 and HO-1 to reduce oxidative stress and apoptosis. This was confirmed using 99mTc-HMPAO brain flow gamma bioassays. Thus, our data suggested GNL as a promising agent for treating neuroinflammation-induced cognitive impairment.

The effect of COVID-19 on nuclear medicine and radiopharmacy activities: A global survey.

Globally, COVID-19 affected radiopharmaceutical laboratories. This study sought to determine the economic, service, and research impacts of COVID-19 on radiopharmacy. This online survey was conducted with the participation of employees from nuclear medicine and radiopharmaceutical companies. The socioeconomic status of the individuals was collected. The study was participated by 145 medical professionals from 25 different countries. From this work, it is evident that 2-deoxy-2-[18F]fluoro-D-glucose (2-[18F]FDG), and 99mTc-labeled macro aggregated albumin 99mTc-MAA were necessary radiopharmaceuticals used by 57% (83/145and 34% (49/145;) respondents, respectively for determining how COVID infections affect a patient's body. The normal scheduling procedure for the radiopharmacy laboratory was reduced by more than half (65%; 94/145). In COVID-19, 70% (102/145) of respondents followed the regulations established by the local departments. Throughout the pandemic, there was a 97% (141/145) decrease in all staffing recruitment efforts. The field of nuclear medicine research, as well as the radiopharmaceutical industry, were both adversely affected by COVID-19.

Asiaticoside Increases Caspase-9 Activity in MCF-7 Cells and Inhibits TNF-α and IL-6 Expression in Nude Mouse Xenografts via the NF-κB Pathway.

Background: We hypothesized that the antitumor effects of asiaticoside on breast cancer are driven by its ability to decrease the expression of tumor inflammation-promoting genes and increase apoptotic signaling. In this study, we aimed to better understand the mechanisms of action of asiaticoside as a chemical modulator or as a chemopreventive agent in breast cancer. Methods: MCF-7 cells were cultured and treated with 0, 20, 40, and 80 µM asiaticoside for 48 h. Fluorometric caspase-9, apoptosis, and gene expression analyses were conducted. For the xenograft experiments, we divided nude mice into the following 5 groups (10 animals per group): group I, control mice; group II, untreated tumor-bearing nude mice; group III, tumor-bearing nude mice treated with asiaticoside at weeks 1-2 and 4-7 and injected with MCF-7 cells at week 3; group IV, tumor-bearing nude mice injected with MCF-7 cells at week 3 and treated with asiaticoside beginning at week 6; and group V, nude mice treated with asiaticoside, as a drug control. After treatment, weight measurements were performed weekly. Tumor growth was determined and analyzed using histology and DNA and RNA isolation. Results: In MCF-7 cells, we found that asiaticoside increased caspase-9 activity. In the xenograft experiment, we found that TNF-α and IL-6 expression decreased (p < 0.001) via the NF-κB pathway. Conclusion: Overall, our data suggest that asiaticoside produces promising effects on tumor growth, progression, and tumor-associated inflammation in MCF-7 cells as well as a nude mouse MCF-7 tumor xenograft model.

Early initiation of insulin attenuates histological and functional changes in the liver of streptozotocin-induced diabetic rats using 99mTc-sulfur colloid functional imaging.

This study aimed to investigate the effect of insulin on the reticuloendothelial system (RES) in the liver and spleen in diabetic rats. Sprague Dawley rats were divided into control, diabetic rats (DM) and diabetic rats treated with insulin (IDM) for 2 weeks. Rats were imaged with technetium-99m-sulfur colloid (99mTc-SC) tracer to determine regional distributions of the tracer for all groups by drawing regions of interest and then obtained the ratios as the cumulative counts of heart, liver, and spleen to the whole body (WB). Liver tissue from sacrificed rats from each group was examined by light and electron microscopy. 99mTc-SC uptake ratios showed a lower liver to WB uptake ratio in the DM rats compared to both controls and IDM rats. Electron microscopy showed severe vacuolization of the hepatocytes of DM rats. The IDM rats show complete resolution of the vacuolization. The early administration of insulin for 2 weeks to diabetic rats could significantly resolve the phagocytic RES function and histological changes in the liver.

Mangiferin protect oxidative stress against deoxynivalenol induced damages through Nrf2 signalling pathways in endothelial cells.

Several cereal grains contain a mycotoxin food contaminant called deoxynivalenol (DON), which presents a significant health risk as it is one of the most commonly found mycotoxins. The current paper examines the ameliorative effect of mangiferin (MAN) in vascular endothelial cells induced through activating the Nrf2 signalling pathway on dietary DON-induced oxidative changes. The study infers that the intercellular reactive oxygen species (ROS) levels and malondialdehyde decrease due to MAN. Other effects include in human umbilical vein endothelial cells (HUVECs), the oxidative stress-induced cell damage is reduced due to protective effects and superoxide dismutase (SOD), and catalase (CAT) activities also reveal an improvement. In HUVECs, the Nrf2-regulated antioxidant enzyme genes' expression is activated by Nrf2 nuclear translocation induction and this activity suppresses the oxidative stress damage. The genes in HUVECs include HO-1 and NQO1. Moreover, in HUVECs, the nucleus translocation of Nrf2 reduces the Nrf2, HO-1, whereas NQO1 expression decreases the cytoprotective effects against oxidative stress reduce with the rejection of Nrf2 with siRNA. This paper pioneers in inferring that oxidative stress-induced HUVECs' cell injury is suppressed by MAN through Nrf2, signalling pathway activation.

Kaempferol Inhibits Zearalenone-Induced Oxidative Stress and Apoptosis via the PI3K/Akt-Mediated Nrf2 Signaling Pathway: In Vitro and In Vivo Studies.

In this study, kaempferol (KFL) shows hepatoprotective activity against zearalenone (ZEA)-induced oxidative stress and its underlying mechanisms in in vitro and in vivo models were investigated. Oxidative stress plays a critical role in the pathophysiology of various hepatic ailments and is normally regulated by reactive oxygen species (ROS). ZEA is a mycotoxin known to exert toxicity via inflammation and ROS accumulation. This study aims to explore the protective role of KFL against ZEA-triggered hepatic injury via the PI3K/Akt-regulated Nrf2 pathway. KFL augmented the phosphorylation of PI3K and Akt, which may stimulate antioxidative and antiapoptotic signaling in hepatic cells. KFL upregulated Nrf2 phosphorylation and the expression of antioxidant genes HO-1 and NQO-1 in a dose-dependent manner under ZEA-induced oxidative stress. Nrf2 knockdown via small-interfering RNA (siRNA) inhibited the KFL-mediated defence against ZEA-induced hepatotoxicity. In vivo studies showed that KFL decreased inflammation and lipid peroxidation and increased H2O2 scavenging and biochemical marker enzyme expression. KFL was able to normalize the expression of liver antioxidant enzymes SOD, CAT and GSH and showed a protective effect against ZEA-induced pathophysiology in the livers of mice. These outcomes demonstrate that KFL possesses notable hepatoprotective roles against ZEA-induced damage in vivo and in vitro. These protective properties of KFL may occur through the stimulation of Nrf2/HO-1 cascades and PI3K/Akt signaling.

Thidiazuron decreases epithelial-mesenchymal transition activity through the NF-kB and PI3K/AKT signalling pathways in breast cancer.

Breast cancer is the major type among the women population globally. The treatment of cancer metastasis has made modest progress due to multiple factors. Thidiazuron (TDZ) is a novel plant growth regulator that has been shown to have anticancer effects. Therefore, we explored the anti-metastatic potentials of TDZ in cell lines by assessing its potential to suppress the epithelial-mesenchymal transition (EMT). We pretreated the BEAS-2B and breast cancer (MDA-MB-231) cells with TDZ and deliberated alteration in a cell viability, mammosphere, migration, NF-кB signalling, PI3K/AKT signalling and matrix metalloproteinase (MMP) expression and analysed the EMT induction by TGF-ß/TNF-α-stimulated BEAS-2B cells. Treatment with TDZ (5-50 µmol) diminished the migration and invasion of the extremely metastatic MDA-MB-231 cells. Additionally, TDZ treatment led to down-regulation of uPAR, uPA, VEGF and MMP-2/-9 expression and up-regulation of TIMP-1/2 expression in these cells. Furthermore, TDZ treatment blocked invasion and EMT in non-tumorigenic BEAS-2B epithelial cells stimulated with TGF-ß/TNF-α.TDZ prevents EMT and may thus block metastasis of breast cancer cells.

Study of the cytotoxicity of asiaticoside on rats and tumour cells.

BACKGROUND: Cancer chemoprevention is considered one of the most promising areas in current cancer research, and asiaticoside, which is derived from the plant Centella asiatica, has a relative lack of systemic toxicity. The purpose of this study was to investigate whether asiaticoside is effective against 7,12-dimethylbenz(a)anthracene (DMBA)-induced carcinogenicity in vitro (MCF-7 and other cells) and in vivo (DMBA-induced rat cancer). METHODS: An MTT assay was performed involving the treatment of MCF-7 cells for 48 h with H2O2 alone and H2O2 + different asiaticoside concentrations. Flow cytometry was performed, and the level of caspase 3, tumour necrosis factor-alpha (TNF-α) and interleukin-1 (IL-1) were quantified. Adult female Sprague-Dawley (SD) rats were divided into five groups designated I (control), II (DMBA-induced cancer), III (pre- and post-treatment with asiaticoside (200 µg/animal) in DMBA-induced cancer), IV (post-treatment with asiaticoside in DMBA-induced cancer), and V (treated with asiaticoside alone, drug control). Twelve weeks post-DMBA, rats developed mammary tumours. Rats either were sacrificed or imaged with MIBI. Histological examination of tumour tissues was performed. Tumour MIBI uptake ratios were determined. The data are expressed as the means ± standard deviation. Appropriate t-test and ANOVA statistical methods were used to compare data. RESULTS: The IC50 of asiaticoside for MCF-7 cells was determined to be 40 µM. Asiaticoside has potential for hydrogen peroxide cytotoxicity, and the caspase-3 activity increased with increasing asiaticoside dose in MCF-7 cells treated for 48 h. The expression of the cytokines TNF-α and IL-1ß was significantly decreased and correlated with MIBI uptake ratios in vitro and in vivo after asiaticoside administration. CONCLUSION: This study demonstrates that asiaticoside is effective in vitro and in vivo in inducing apoptosis and enhancing anti-tumour activity.

Choline treatment affects the liver reticuloendothelial system and plasma fatty acid composition in diabetic rats.

PURPOSE: This study investigated effects of choline treatment on hepatic reticuloendothelial and biliary functions and plasma fatty acid composition in diabetic rats. METHODS: Diabetes was induced by streptozotocin (STZ). Choline was administered to untreated rats and a portion of STZ-treated rats for two sequences of five consecutive days, separated by a 2-day interval. Hepatic functions were studied using (99m) Tc Tin (II) colloid (TIN) and 99 mTc mebrofenin [bromo-iminodiacetic acid (BrIDA)] imaging. The TIN-uptake ratios (organ/whole body) of heart, liver and spleen, and the BrIDA-uptake ratios (organ or tissue/whole body) of liver, biliary tree and abdomen were obtained following imaging studies. Fatty acids were analysed by GC/MS. RESULTS: Choline treatment did not attenuate hyperglycaemic development. Diabetic rats showed (i) a decreased TIN-uptake ratio in liver with co-increased ratios in heart and spleen; choline treatment diminished these changes, (ii) elevated BrIDA-uptake ratios in biliary tree and abdomen but not in liver; choline treatment did not attenuate the elevations and (iii) decreases in plasma palmitoleic acid and oleic acid, reflecting an impaired stearoyl-CoA desaturase function; choline treatment did not affect the diminutions, but caused a decrease in arachidonic acid with a co-increase in linoleic acid. Some rats developed hypoproteinemia (HPO). HPO rats also exhibited decreases in plasma palmitoleic acid and oleic acid. Diabetes caused almost absence of palmitoleic acid in HPO rats. Choline treatment exerted no effect on the plasma fatty acid composition of diabetic HPO rats. CONCLUSIONS: Choline treatment affected hepatic reticuloendothelial function and plasma fatty acid composition, but not hepatobiliary function, in diabetic rats. Whether choline treatment is beneficial requires further studies.

Assessment of tracer 99mTc(V)-DMSA uptake as a measure of tumor cell proliferation in vitro.

PURPOSE: To examine whether (99m)Tc(V)-DMSA could be used as a non-invasive measure of cancer cell proliferation. METHODS: Human breast cancer MCF-7, MDA-MB-231 and pII, and prostate cancer PC-3 cell lines were grown to 30, 50 and 100% confluency and pulsed with (99m)Tc(V)-DMSA in media for 60 min at 37°C. DNA synthesis was analysed by quantification of the S phase using flow cytometry, [methyl-(3)H]thymidine incorporation and expression of proliferation markers PCNA and Ki-67 using realtime PCR. One way ANOVA was used to compare groups. RESULTS: In all cell lines rates of (99m)Tc(V)-DMSA uptake were inversely related to cell density. This was paralleled by similar trends in S phase proportions, [methyl-(3)H]thymidine incorporation and expression of PCNA and Ki-67. CONCLUSION: Rates of (99m)Tc(V)-DMSA uptake into different types of tumour cells correlate well with cell density that is useful as a non-invasive measure of tumour cellular proliferation in vivo.

Effect of asiaticoside on 99mTc-tetrofosmin and 99mTc-sestamibi uptake in MCF-7 cells.

UNLABELLED: This study was done to examine the effect of asiaticoside on MCF-7 cell uptake of (99m)Tc-tetrofosmin ((99m)Tc-Tfos) and (99m)Tc-sestamibi ((99m)Tc-MIBI). METHODS: The 3-(4,5-dimethylthiozol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was used to evaluate the effect of a 50% inhibitory concentration of asiaticoside on MCF-7 cell proliferation. MCF-7 cells were treated with 10, 20, 30, 40, and 50 µM asiaticoside for 48 h and then incubated with 59.2 MBq of either (99m)Tc-Tfos or (99m)Tc-MIBI tracer for 60 min. The uptake of the tracers was measured with a dose calibrator. RESULTS: The 50% inhibitory concentration of asiaticoside for MCF-7 cells was determined with the MTT assay to be 40 µM. The uptake results were expressed as the mean ± SE radioactivity in MBq/mg of protein, and P values were also calculated (P values of 0.03 indicated significant differences). In the control (no asiaticoside) and at 10, 20, 30, 40, and 50 µM asiaticoside, the mean levels of (99m)Tc-Tfos uptake were 0.79 (SE, 0.059) (P = 0.14), 0.84 (SE, 0.057) (P = 0.60), 0.47 (SE, 0.034) (P = 0.03), 0.40 (SE, 0.050) (P = 0.03), 0.37 (SE, 0.050) (P = 0.03), and 0.15 (SE, 0.023) (P = 0.03), respectively; the mean levels of (99m)Tc-MIBI uptake were 0.95 (SE, 0.007) (P = 0.14), 0.81 (SE, 0.009) (P = 0.60), 0.79 (SE, 0.019) (P = 0.03), 0.63 (SE, 0.004) (P = 0.03), 0.13 (SE, 0.006) (P = 0.03), and 0.07 (SE, 0.008) (P = 0.03), respectively. Asiaticoside concentrations of 10, 20, 30, 40, and 50 µM revealed the uptake kinetics for both (99m)Tc-Tfos and (99m)Tc-MIBI in MCF-7 cells. (99m)Tc-Tfos and (99m)Tc-MIBI showed similar trends; the radioactivity uptake was dose dependent, and asiaticoside inhibited 16% and 47% of (99m)Tc-Tfos uptake and (99m)Tc-MIBI uptake in MCF-7 cells, respectively. CONCLUSION: This study showed that asiaticoside, acting as a biochemical modulator, may induce apoptosis and enhance antitumor activity in MCF-7 cells, as determined by (99m)Tc-Tfos and (99m)Tc-MIBI uptake. These findings are promising for cancer chemotherapy. Future studies should be performed to confirm our findings and to further delineate the clinical role of asiaticoside.

(99m)Tc sulfur colloid and (99m)Tc mebrofenin hepatobiliary functional liver imaging in normal and diabetic rats.

OBJECTIVES: To use (99m)Tc sulfur colloid ((99m)Tc-SC) and (99m)Tc mebrofenin ((99m)Tc-BrIDA) to study liver function in normal and diabetic rats. MATERIALS AND METHODS: Radionuclide imaging was performed on 2 groups of rats, using (99m)Tc-SC for one group and (99m)Tc-BrIDA for the other (20 rats per group) before and after induction of diabetes mellitus (DM) using streptozotocin administration (55 mg/kg i.p.). Dynamic acquisition was obtained for 1 h after the injection of 37 MBq of radiotracer. For the (99m)Tc-SC group, organ/tissue uptake was determined by drawing regions of interest (ROI) over the heart, liver, spleen and also the whole body (WB). The ratio of the ROI of each organ to the WB ROI was calculated. For (99m)Tc-BrIDA, ratios of cumulative count rates in liver, liver parenchyma, biliary tree and abdomen ROI to a WB ROI were also calculated. Statistical analysis was performed to compare the ratios of organ/tissue uptake to WB uptake before and after DM induction using the paired t test. RESULTS: (99m)Tc-SC uptake ratios (means ±SD) showed a lower liver-to-WB uptake ratio (0.75 ± 0.05) in the rats after DM induction compared to baseline (0.81 ± 0.06), while the cardiac blood pool showed higher uptake ratios in the rats after DM induction (p = 0.026). For (99m)Tc-BrIDA, there was no significant difference in radiotracer uptake ratios obtained from the rats before and after DM induction (p = 0.41). CONCLUSION: Using functional liver imaging, there was a statistically significant decrease in the liver phagocytic/reticuloendothelial system function after DM induction, as evidenced by decreased (99m)Tc-SC liver uptake and increased blood pool compared to prediabetes, while the hepatobiliary function remained unchanged after DM induction using (99m)Tc-BrIDA imaging.

Intracellular localization of (99m)Tc-d,l-HMPAO in rat blood.

OBJECTIVE: The objective of this study was to investigate the intracellular localization of technetium 99m-hexamethylpropylene amine oxime ((99m)Tc-d,l-HMPAO) in rat red blood cells (RBCs) and plasma. MATERIALS AND METHODS: Sprague-Dawley rats (n = 36) were injected with 129.5 MBq (99m)Tc-d,l-HMPAO. Using a heparinized syringe, 2-ml blood samples were withdrawn from the rats 10, 20, 30, 40, 50 and 60 min after (99m)Tc-d,l-HMPAO injection. Blood was centrifuged for 10 min at 2,240 rpm to separate RBCs from plasma. A chloroform/methanol/distilled water mixture was added to the RBCs and plasma in separate tubes to extract (99m)Tc-d,l- HMPAO into aqueous (Aq) and lipid (Lp) phases. The (99m)Tc-d,l-HMPAO activities in Aq and Lp phases were measured separately using a dose calibrator. The data were analyzed using Student's t test and a one-way ANOVA. RESULTS: More than 70% of the injected dose of (99m)Tc-d,l-HMPAO was retained in the RBCs for up to 60 min, compared to less than 20% in plasma. The uptakes in the Aq for plasma were 88.77 +/- 1.04, 94.23 +/- 2.42, 90.15 +/- 1.88, 98.59 +/- 0.89, 96.42 +/- 1.50 and 97.59 +/- 1.92% at 10, 20, 30, 40, 50 and 60 min after injection, respectively. The corresponding uptakes in the Lp phases were: 11.23 +/- 0.04, 5.77 +/- 0.07, 9.85 +/- 0.06, 1.41 +/- 0.08, 3.58 +/- 0.04 and 2.41 +/- 0.06, respectively. Similar values (% +/- SE) and trends were obtained in the Aq and Lp phases for RBCs. A highly significant difference between the Aq and Lp phases at each time point studied was observed (p < 0.0001). There was no statistically significant difference between the time points in the Aq phase, but there was a significant difference (p < 0.001) between time points in the Lp phase. CONCLUSION: This study demonstrated that most of the intracellular localization of (99m)Tc-d,l-HMPAO was in the Aq phase compared to Lp phase in both RBCs and plasma up to 60 min.

Perfusion scanning using 99mTc-HMPAO detects early cerebrovascular changes in the diabetic rat.

BACKGROUND: 99mTc-HMPAO is a well-established isotope useful in the detection of regional cerebral blood flow. Diabetes gives rise to arterial atherosclerotic changes that can lead to significant end organ dysfunction, prominently affecting perfusion to the heart, kidneys, eyes and brain. In the current study, we investigated the role of 99mTc-HMPAO cerebral perfusion scans in detecting early vascular changes in the diabetic brain. METHODS: Cerebral perfusion studies were performed on both control and streptozotocin-(STZ) induced diabetic male Wistar rats. Rat brain imaging using a gamma camera was performed for each group 0.5, 2, 4, and 24 hours post 99mTc-HMPAO injection. Data processing for each cerebral perfusion scan was performed by drawing a region of interest (ROI) circumferentially around the brain (B). Background (BKG) due to signal from the soft tissue of each rat was subtracted. Brain 99mTc-HMPAO uptake minus background counts (net brain counts; NBC) were then compared between the two groups. RESULTS: The NBC (mean +/- SD) for the STZ group were statistically significantly higher (p = 0.0004) than those of the control group at each of the time points studied. CONCLUSION: 99mTc-HMPAO brain scan may be useful in the detection of early atherosclerotic changes in the diabetic rat brain.

[Methyl-3H]-choline incorporation into MCF-7 cells: correlation with proliferation, choline kinase and phospholipase D assay.

BACKGROUND: [Methyl-3H]-choline is a promising new positron emission tomography (PET) agent used for cancer imaging whose mechanism has still not fully been elucidated. In this study, whether [methyl-3H]-choline determined by measuring the activity of choline kinase (ChoK) and phospholipase D (PLD) in rapidly proliferating and confluent breast cancer MCF-7 cells is related to cell proliferation or not was investigated. MATERIALS AND METHODS: The activity of ChoK and PLD were determined using ion exchange chromatography and transphosphatidylation assay respectively. RESULTS: [Methyl-3H]-PCho content expressed as pmol mg(-1) protein(-1) min(-1) (n = 6) was significantly higher in the exponentially growing (484.04 +/-2 0.23) compared with confluent (70.35 +/-9.83) cells using Student's t-test (p < 0.001). Moreover, PLD activity expressed as the mean (n = 6) (disintegration per minute (d.p.m.)/microg protein +/- SD (mean S phase +/- SD)) showed significantly higher (p < 0.001) activity in the exponentially growing cells (196.39 +/- 2.21 d.p.m./microg protein (39.69 +/- 4.00%)) compared with confluent cells (99.10 +/- 1.35 d.p.m./microg protein (9.33 +/- 0.82%)). CONCLUSION: This study indicates that the major water-soluble choline metabolite was phosphocholine (PCho) as a consequence of increased ChoK and PLD activity in the exponentially growing cells compared to confluent cells.

Effects of flutamide on [methyl-(3)h]-choline uptake in human prostate cancer-3 cells: a pilot study.

BACKGROUND: Positron emission tomography using [methyl-(11)C]-choline is effective in imaging many types of cancer, especially prostate cancer (PC). The antiandrogen flutamide is often used as part of the initial treatment of PC. Data on the effect of flutamide on and methylcholine incorporation into PC-3 cells are lacking in the experimental and literature work. OBJECTIVES: The aims of this study were to assess whether human PC-3 cells are susceptible to flutamide and whether the drug modulates the uptake of [methyl-(3)H]-choline into these cells. METHODS: PC-3 cells were treated for 3 days with flutamide (≤100 nmol/L), inhibiting growth by 20% to 70% with control cells included. Two viability tests (cytotoxic analyses), the thiazole blue assay and the trypan blue exclusion method, were used to determine the median inhibitory concentration for flutamide (10 nmol/L). Control and flutamide-treated cells were incubated with [methyl-(3)H]-choline for 10 minutes and then in nonradioactive medium for 10 minutes to simulate the rapid blood clearance of [methyl-(11)C]-choline tracer that occurs within 5 to 20 minutes, and then extracted using organic and aqueous solvents to determine the intracellular distribution of the tracer. Protein assay and flow-cytometry analysis were used to determine protein content and DNA synthesis in both control and treated cells. The uptake of [methyl-(3)H]-choline was normalized to protein content and expressed as mean (SD) dpm/1Jg protein (n = 6). RESULTS: PC-3 cell proliferation was inhibited with flutamide treatment. After treatment of PC-3 cells with flutamide 10 nmol/L for 3 days, cells accumulated DNA during the S phase. Mean (SD) [methyl-(3)H]-choline uptake was found to be significantly lower with flutamide 10-nmol/L-treated cells compared with control cells (65.95 [0.72] vs 114.21 [0.57] dpm/1Jg protein; P < 0.001); the difference between the 5-nmol/L-treated cells and controls was nonsignificant. CONCLUSIONS: In this pilot study, flutamide inhibited tumor cell growth and proliferation and decreased (modulated) the uptake of [methyl-(3)H]-choline into androgen receptor-negative PC-3 cells. These results suggest that flutamide might inhibit proliferation by an androgen-independent mechanism.

Role of 99mTc-(V)DMSA in detecting tumor cell proliferation.

Pentavalent technetium-99m dimercaptosuccinic acid ((99m)Tc-(V)DMSA) is a tumor-seeking agent which was introduced to evaluate, image, and manage many types of cancers. In this review, the beginning of, and the most recent applications of using this agent was appraised. The relation with tumor cell detection and proliferation was reported and several mechanisms of uptake of (99m)Tc-(V)DMSA in tumor cells are described.

[methyl-3H]Choline incorporation into MCF7 tumour cells: correlation with proliferation.

PURPOSE: The aim of the study was to investigate the intracellular location of [methyl-(3)H]choline in MCF7 tumour cells and to determine the relationship between [methyl-(3)H]choline incorporation and proliferation. METHODS: Tumour cells were incubated with [methyl-(3)H]choline for 10 min, and then in cold medium to simulate the rapid blood clearance of [methyl-(11)C]choline. Labelled metabolites were then extracted from cells by treating them with organic and aqueous solvents to determine the distribution of tracer between phospholipid and water-soluble metabolite pools. Aqueous extracts were subjected to thin-layer chromatography, ion exchange chromatography and a choline extraction procedure to identify (3)H-containing metabolites. Procedures were carried out on fast- and slow-growing populations of MCF7 cells to determine the relationship between choline incorporation and proliferation. RESULTS: Only about 5% of [methyl-(3)H]choline was present as phospholipid. [methyl-(3)H]choline incorporation was found to be related to S-phase fraction. In another experiment, [methyl-(14)C]choline incorporation was found to be correlated with [methyl-(3)H]thymidine incorporation. The V(max) of choline uptake was found to be increased whilst K(m) was decreased in populations of MCF7 cells with higher proliferative fractions, compared with populations having lower proliferative fractions. CONCLUSION: Choline incorporation into tumour cells under conditions that simulate rapid blood clearance of [methyl-(11)C]choline is correlated with proliferation. Most of the activity (about 95%) was in the non-lipid fraction of the cell.