Enzymatically-crosslinked gelatin hydrogels containing paenipeptin and clarithromycin against carbapenem-resistant pathogen in murine skin wound infection.

BACKGROUND: The recent rise and spread of carbapenem-resistant pathogens pose an urgent threat to public health and has fueled the search for new therapies. Localized delivery of topical antibiotics is an alternative for the treatment of infected wounds caused by drug-resistant pathogens. In this study, we aimed to develop antimicrobial-loaded hydrogels for topical treatment of wound infections in a murine skin wound infection. RESULTS: Paenipeptin analogue 1, a linear lipopeptide, potentiated clarithromycin against multidrug-resistant Acinetobacter baumannii, Enterobacter cloacae, Escherichia coli, and Klebsiella pneumoniae. Enzymatically-crosslinked gelatin hydrogels were developed to encapsulate paenipeptin analogue 1 and clarithromycin. The encapsulated antimicrobials were gradually released from hydrogels during incubation, reaching 75.43 and 53.66% for paenipeptin and clarithromycin, respectively, at 24 h. The antimicrobial-loaded hydrogels containing paenipeptin and clarithromycin synergistically resulted in 5-log reduction in carbapenem-resistant A. baumannii within 6 h in vitro. Moreover, the antimicrobial-loaded hydrogels reduced 3.6- and 2.5-log of carbapenem-resistant A. baumannii when treated at 4 or 20 h post infection, respectively, in a murine skin wound infection. CONCLUSIONS: Enzymatically-crosslinked gelatin hydrogels loaded with paenipeptin analogue 1 and clarithromycin exhibited potent therapeutic efficacy against carbapenem-resistant A. baumannii in murine skin wound infection.

Paenipeptin Analogues Potentiate Clarithromycin and Rifampin against mcr-1-Mediated Polymyxin-Resistant Escherichia coli In Vivo.

Polymyxin resistance mediated by the mcr-1 gene threatens the last-resort antibiotics. Linear lipopeptide paenipeptin analogues 1 and 15 disrupted the outer membrane of Gram-negative pathogens and potentiated clarithromycin and rifampin against mcr-1-positive Escherichia coli from the FDA-CDC Antimicrobial Resistance Isolate Bank. In the presence of paenipeptin, clarithromycin and rifampin resulted in over 3-log reduction of E. coliin vitro Moreover, paenipeptin-antibiotic combinations significantly reduced E. coli in a murine thigh infection model.

Proteomic profiling of tear fluid as a promising non-invasive screening test for colon cancer.

BACKGROUND: Current screening options for colorectal cancer (CRC) are either invasive (colonoscopy) or have lower sensitivity to identify pre-malignant lesions (fecal immunochemical test). We proposed to identify protein profiles in tears of patients with both pre-malignant polyps and CRC; these profiles could have potential as a noninvasive screening test. METHOD: Colonoscopy patients were divided into "high risk" group (CRC and tubular adenomatous polyp) and "low risk" (normal and hyperplastic polyps). Tear fluids from patients were analyzed by Liquid Chromatography Mass Spectrometry/Mass Spectrometry. The data were analyzed for protein expression, protein-protein interaction and gene set enrichment. RESULTS: The results showed 80 proteins (18 up-regulated and 62 down-regulated) significantly differentiated in "high-risk" compared to "low-risk"; Twenty-eight of these show protein-protein interactions, 9 of which were associated with pathways demonstrated to be altered in CRC patients. CONCLUSION: Our pilot data, though limited, demonstrated tear protein profiling could distinguish the groups of patients with and without colon lesions.

Oral glutamine protects against acute doxorubicin-induced cardiotoxicity of tumor-bearing rats.

Doxorubicin (DOX), a widely used anticancer drug, has a dose-dependent cardiotoxicity, attributed mainly to free radical formation. The cardiomyocyte oxidative stress occurs rapidly after DOX treatment, resulting in harmful modifications to proteins, lipids, and DNA. Previous data showed that oral l-glutamine (Gln) prevented cardiac lipid peroxidation and maintained normal cardiac glutathione (GSH) levels in DOX-treated rats. Our aim in this study was to examine the effect of Gln on DOX-induced cardiac oxidative stress in a tumor-bearing host. Female Fisher344 rats with implanted MatBIII mammary tumors were randomized into 2 groups: a Gln group that received l-Gln (1 g.kg(-1).d(-1)) (n = 10) via a Gln-enriched diet and/or gavage with 50% Gln suspension during the whole experiment and a control group that was fed the same diet formulation without Gln and/or were gavaged with water. All rats received a single injection of 12 mg/kg DOX and were killed 3 d later. GSH levels of hearts, livers, tumors, and blood, as well as cardiac histological alterations, lipid peroxidation, peroxinitrite levels, and caspase-3 activation were determined. Cardiac physiologic alterations were assessed by ultrasound imaging before and 3 d after DOX administration. The Gln supplementation resulted in lower cardiac lipid peroxidation and peroxintrite levels and elevated cardiac catalase enzyme activity and GSH compared with the controls, without affecting those of the tumors. DOX-induced alterations of the echocardiographic parameters were significantly reduced in the Gln-supplemented rats. These data indicate that Gln is able to reduce the oxidative damage of cardiomyocytes that occurs soon after DOX administration and thus protects the heart of a tumor-bearing host from DOX-induced cardiomyopathy.

Dantrolene Attenuates Cardiotoxicity of Doxorubicin Without Reducing its Antitumor Efficacy in a Breast Cancer Model.

Dysregulation of calcium homeostasis is a major mechanism of doxorubicin (DOX)-induced cardiotoxicity. Treatment with DOX causes activation of sarcoplasmic reticulum (SR) ryanodine receptor (RYR) and rapid release of Ca2+ in the cytoplasm resulting in depression of myocardial function. The aim of this study was to examine the effect of dantrolene (DNT) a RYR blocker on both the cardiotoxicity and antitumor activity of DOX in a rat model of breast cancer. Female F344 rats with implanted MAT B III breast cancer cells were randomized to receive intraperitoneal DOX twice per week (12 mg/kg total dose), 5 mg/kg/day oral DNT or a combination of DOX + DNT for 3 weeks. Echocardiography and blood troponin I levels were used to measure myocardial injury. Hearts and tumors were evaluated for histopathological alterations. Blood glutathione was assessed as a measure of oxidative stress. The results showed that DNT improved DOX-induced alterations in the echocardiographic parameters by 50%. Histopathologic analysis of hearts showed reduced DOX induced cardiotoxicity in the group treated with DOX + DNT as shown by reduced interstitial edema, cytoplasmic vacuolization, and myofibrillar disruption, compared with DOX-only-treated hearts. Rats treated with DNT lost less body weight, had higher blood GSH levels and lower troponin I levels than DOX-treated rats. These data indicate that DNT is able to provide protection against DOX cardiotoxicity without reducing its antitumor activity. Further studies are needed to determine the optimal dosing of DNT and DOX in a tumor-bearing host.

Dead or alive? Autofluorescence distinguishes heat-fixed from viable cells.

PURPOSE: A proof-of-concept study to evaluate a new autofluorescence method to differentiate necrotic thermally fixed cells from viable tissue following thermal ablation. METHODS: A conductive interstitial thermal therapy (CITT) device was used to ablate swine mammary tissue and rabbit VX-2 carcinomas in vivo. The ablated regions and 10-mm margins were resected 24 h following treatment, embedded in HistOmer and sectioned at 3 mm. The fresh sections were evaluated for gross viability with triphenyl tetrazolium chloride, 1 h post-resection. Representative non-viable and viable areas were then processed and embedded into paraffin, and sectioned at 5 microm. Standard H&E staining and proliferating cell nuclear antigen (PCNA) immunohistochemistry were compared against autofluorescence intensity, at 488-nm wavelength, for cellular viability. RESULTS: Heat-fixed cells in non-viable regions exhibit increased autofluorescence intensity compared to viable tissue (area under receiver operating characteristics (ROC) curve = 0.96; Mann-Whitney P < 0.0001). An autofluorescence intensity-based classification rule achieved 92% sensitivity with 100% specificity for distinguishing non-viable from viable samples. In contrast, PCNA staining did not reliably distinguish heat-fixed, dead cells from viable cells. CONCLUSIONS: Examination of H&E-stained sections using autofluorescence intensity-based classification is a reliable and readily available method to accurately identify heat-fixed cells in ablated surgical margins.

Conductive interstitial thermal therapy (CITT) inhibits recurrence and metastasis in rabbit VX2 carcinoma model.

PURPOSE: To investigate the potential of conductive interstitial thermal therapy (CITT) to inhibit recurrence and metastasis in a partially resected tumour model. METHOD: Fifteen New Zealand white rabbits were implanted with VX2 tumour intramuscularly in the rear thigh. Once the tumour size reached 20-25 mm in diameter, three animals were randomly selected to serve as controls, while the remaining animals were designated as the study group and treated with CITT. In the CITT group, the partially resected tumour and margins were thermally ablated. In the control group the tumour was partially resected to simulate positive margins. The animals were monitored for up to 12 weeks. At the endpoint, the animals were sacrificed, and whole-body diagnostic necropsy was conducted immediately. RESULTS: Recurrences and metastatic lesions were observed in iliac and popliteal lymph nodes and abdomens of all control animals. In contrast, the observed rate of recurrence and metastatic lesion was 0% among CITT-treated animals, significantly less than the >or=50% null-hypothesis rate expected upon treatment failure (exact binomial P = 0.0002). Complete histopathological healing was obtained in 2 of 12 rabbits, and residual inflammation remained at the ablation site up to 12 weeks post-ablation in 10 of 12 rabbits. This pattern of necrosis and inflammatory response was not observed in any of the control rabbits. CONCLUSIONS: The CITT device effectively ablated partially resected VX2 carcinoma in a rabbit model, and inhibited recurrence and metastasis in this model. CITT evoked an inflammatory response that may be linked to the mechanism involved in reduced metastatic spread.

Glutamine affects glutathione recycling enzymes in a DMBA-induced breast cancer model.

Malignancy depletes host glutathione (GSH) levels to increase treatment-related toxicity and increases itself to resist the treatments. Our previous studies have shown that dietary glutamine (GLN) prevented 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors through enhancing gut GSH release and reducing tumor GSH level. In addition, GSH synthesis, metabolism, and recycling are accomplished in gamma-glutamyl cycle. We hypothesized that the GLN prevention might be through a differential regulation of the gamma-glutamyl cycle enzymes. Female Sprague-Dawley rats were randomized into DMBA-tumor bearing, DMBA-treated, and control groups subdivided into GLN and water groups. GLN supplementation was given at 1 g/kg/day by gastric gavage. The activities and messenger RNA levels of gamma-glutamyl transpeptidase (GTP), gamma-glutamylcysteine synthetase (GCS), 5-oxo-L-prolinase (OPase), gamma-glutamyl transferase (GTF), and glutaminase (GLNase) were determined in gut mucosa and breast tumor using specific enzyme assays and semiquantitative reverse transcription polymerase chain reaction. GLN upregulated gut GTP, GCS, OPase, and GLNase in DMBA-tumor bearing, DMBA-treated, and/or control rats; however, it downregulated these enzymes in the tumor. The paradoxical effect of GLN on key GSH recycling enzymes in the gut versus tumor suggests that dietary supplemental GLN could be used in the clinical practice to increase the therapeutic index of cancer treatments by protecting normal tissues from, and sensitizing tumor cells to, chemotherapy and radiation-related injury.

Oral glutamine prevents DMBA-induced mammary carcinogenesis via upregulation of glutathione production.

OBJECTIVE: Glutamine is suggested to participate in glutathione synthesis. Furthermore, there is a positive relation between glutathione level and natural killer (NK) cell activity. Previously we demonstrated that supplemental glutamine inhibited tumor growth in an implantable tumor model and 7,12-dimethylbenz[a]anthracene (DMBA)-induced oral squamous cell carcinoma model; these reductions were associated with enhancing NK cell cytotoxicity, blood glutathione levels, and/or gut glutathione release. Therefore, we hypothesized that oral glutamine might suppress DMBA-induced mammary carcinogenesis by upregulation of glutathione production and/or augmentation of NK cell activity. METHODS: Rats were randomized to six groups: DMBA + glutamine, DMBA + Freamine, DMBA + water, oil + glutamine, oil + Freamine, or oil + water. At age 50 d, rats were gavaged with DMBA or sesame oil. Rats also received glutamine, Freamine, or water by gavage from 1 wk before DMBA administration until sacrifice at 1, 2, 4, or 11 wk after DMBA. Tumor appearance, blood, gut mucosa and breast glutamine and/or glutathione concentrations, and NK cell cytotoxicity were measured. The gut extractions, defined as the difference of concentrations across the gut, were calculated. RESULTS: Oral glutamine reduced the tumorigenesis of DMBA by 50% in this model. DMBA altered the difference of glutathione concentrations across the gut; however, oral glutamine maintained the normal gut glutathione release. NK cell activities were lower in DMBA groups at early (week 1) and late (week 11) time points, but oral glutamine recovered the NK cell activity only at week 11. In addition, blood, gut, and breast glutathione concentrations were enhanced by glutamine supplementation. CONCLUSION: These results indicate that one of the mechanisms of dietary glutamine anticancer action is through upregulating gut glutathione metabolism.

Conductive interstitial thermal therapy (CITT) device evaluation in VX2 rabbit model.

We have developed a conductive interstitial thermal therapy (CITT) device to precisely and reliably deliver controlled thermal doses to the surgical margins at the cavity site following tumor resection, intraoperatively. The temperature field created by CITT ablation of a perfused tissue was modeled with a finite element package Femlab. The modeling suggested that a maximum probe temperature of 120 degrees C and an ablation time of 20 minutes were required to ablate highly perfused tissue such as the VX2 carcinoma. Deployable pins enable faster and more reliable thermal ablation. The model predictions were tested by thermal ablation of VX2 carcinoma tumors implanted in adult New Zealand rabbits. The size of the ablated region was confirmed with a viability stain, triphenyltetrazolium chloride (TTC). Histopathological examination revealed 3 regions in the ablated area: a carbonized region (1-3 mm); a region that contained thermally fixed cells; and an area of coagulated necrosis cells. Cells in the thermally fixed region stained for PCNA (proliferating cell nuclear antigen) and were bounded by the carbonized layer at the cavity wall, and by necrotic cells that exhibit nuclear fragmentation and cell dissociation, 5 to 10 mm away from the CITT probe. Adjacent tissue outside the target region was spared with a clear demarcation between ablated and normal viable tissue. It is suggested that the CITT device can be used, clinically, to inhibit local recurrence by creating negative surgical margins following the resection of a primary tumor in non-metastatic early staged tumors.

Immunotargeted photodynamic therapy for cholesteatoma: in vitro results with anti-EGFR-coated indocyanine green nanocapsules.

HYPOTHESIS: The objective was to test the hypothesis that immunotargeted photodynamic therapy (IT-PDT) using anti-epithelial growth factor receptor (EGFR)-coated indocyanine green (ICG) nanocapsules would selectively kill cholesteatoma-derived keratinocytes while sparing middle ear-derived mucosa cells in vitro. BACKGROUND: Rates of residual cholesteatoma caused by incomplete microsurgical removal are unacceptably high; thus, development of an adjuvant therapy to safely destroy undetected residual cholesteatoma cells would be desirable. IT-PDT is a possible means to achieve this end. METHODS: ICG nanocapsules coated with anti-EGFR were synthesized and applied to cholesteatoma-derived keratinocytes and middle ear mucosa cells in vitro. Selective binding to keratinocytes was evaluated by fluorescence microscopy. Activation of ICG was undertaken by applying near-infrared light (810 nm) at an applied energy dose of 1,080 J/cm. Cell death was evaluated 2 hours after treatment with trypan blue staining. RESULTS: Selective and robust nanocapsule binding to keratinocytes, but not mucosa cells, was confirmed by preapplication and postapplication fluorescence measurements. A keratinocyte cell death rate of 70.12% ± 2.50% was achieved, whereas negligible mucosa cell death was observed. Negligible cell death was also observed for both cell types with application of the nanocapsules alone or with application of near-infrared light alone. CONCLUSION: Anti-EGFR ICG nanocapsules applied topically and activated as part of an IT-PDT scheme results in a high rate of cholesteatoma-derived keratinocyte cell death while negligibly affecting middle ear mucosal cells in vitro. These preliminary findings suggest that this is a feasible concept and that further investigation is warranted.

Oral glutamine (AES-14) supplementation inhibits PI-3k/Akt signaling in experimental breast cancer.

BACKGROUND: 7,12-dimethylbenz [a] anthracene (DMBA) administration to pubertal rats causes breast tumors and inhibits glutathione (GSH) production. Our previous results have established that oral glutamine (GLN) supplementation significantly reduced tumor development, restored the depressed GSH production, and caused a significant decrease in the circulating levels of insulinlike growth factor-1 (IGF-1). The present study was designed to investigate the involvement of the IGF-1-activated phosphatidylinositol 3 kinase (PI-3K)/Akt apoptotic signaling pathway. MATERIALS AND METHODS: Forty female Sprague-Dawley rats were randomly divided into 4 groups: DMBA+GLN (n = 16), DMBA+water (n = 8), Oil+GLN (n = 8) and Oil+water (n = 8). At the age of 50 days, rats received a single dose of 100 mg/kg DMBA (n = 24) or sesame oil (n = 16) and were gavaged with a GLN suspension formulation (AES-14) or water for the duration of the entire experiment. The animals were killed 11 weeks after the DMBA application, and the levels of IGF-1, IGF-1 receptor (IGF-IR), Akt, Bcl-2 and Bad in tumorous and nontumorous breast tissue samples were measured by Western blot analysis. RESULTS: GLN supplementation resulted in a significant decrease in the levels of IGF-1, IGF-IR, Akt, and Bcl-2 in nontumorous samples. At the same time, the levels of pro-apoptotic protein Bad were significantly elevated. The samples collected from tumor tissues showed lower levels of IGF-1, Akt, Bcl-2, Bad, and IGF-IR in comparison with nontumorous tissues. CONCLUSIONS: GLN supplementation inhibited the PI-3K/Akt pathway that is thought to be important in increasing cell survival during tumorigenesis. These results are in agreement with our hypothesis that GLN counteracts the effects of DMBA and blocks carcinogenesis in vivo.

Effect of glutamine on the initiation and promotion phases of DMBA-induced mammary tumor development.

BACKGROUND: Oral glutamine (GLN) has been shown to up-regulate tissue glutathione (GSH), augment natural killer (NK) cell activity, and prevent tumor growth in an implantable breast cancer model (MTF-7). We hypothesized that dietary GLN would likewise antagonize the induction or promotion of tumor formation by 7,12-dimethylbenz[a]anthracene (DMBA) via up-regulation of GSH or augmentation of NK activity. METHODS: At age 55 days, 81 Sprague-Dawley rats were gavaged with a one-time dose of 80 mg/kg DMBA, time 0. Rats were randomized into 3 groups (GLN+DMBA, Freamine [FA]+DMBA, water (H2O)+DMBA), pair-fed chow, and gavaged with 1.0 g/kg/day GLN or isonitrogenous amount of FA or H2O for the indicated times: PreFed (-1 to + 16 weeks), Short-Fed (-1 to + 1 weeks) and PostFed (+ 1 to +16 weeks). After 16 weeks, rats were killed and examined for mammary tumors, blood was assayed for GLN and GSH content, and spleens were assayed for NK cytotoxicity. RESULTS: Over the 4-month study period, there was no significant difference in tumorigenesis between FA and H2O groups, regardless of timing of feeding and amino acid diet, except GLN. In Pre- and PostFed GLN groups, there was no significant difference between groups, but there were significant decreases in tumorigenesis in GLN groups compared with either FA or H2O groups. However, in the Short-Fed group, there was no significant difference in tumorigenesis from the GLN, FA, or H2O groups. CONCLUSIONS: Continuously supplemented GLN significantly reduced DMBA-induced breast cancer growth when compared with the non-GLN-supplemented and Short-Fed supplemental GLN groups. Furthermore, GLN appears to have its primary effect on promotion and not initiation of tumor formation. This decreased tumor formation was associated with significantly higher arterial GLN and GSH levels and NK activity at killing in the GLN+DMBA group. Protein in the presentation of FA did not promote or prevent tumor growth. These data indicate that GLN may be useful in the chemoprevention of breast cancer.

Oral glutamine reduces radiation morbidity in breast conservation surgery.

This study examined the effect of oral glutamine (Gln) on radiation injury in breast cancer patients undergoing radiation therapy. The radiation injury was evaluated using Radiation Therapy Oncology Group (RTOG) scales. Cosmesis was scored. Blood Gln and glutathione (GSH) levels were determined. Serum protein profiling was determined using surface-enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS). Patients receiving Gln scored significantly better in RTOG score than the patients receiving placebo. Cosmetic scores averaged excellent in the Gln group vs fair to good in the placebo group. Blood Gln and GSH levels were significantly higher in the Gln group vs the placebo group. Serum protein profiling with SELDI-TOF MS identified a novel Gln-responsive protein that showed amino acid similarity with myoglobin. These results suggest that Gln is an effective way to reduce radiation morbidity to breast cancer and is associated with the increased expression of a novel serum protein biomarker.

Increased efficacy and reduced cardiotoxicity of metronomic treatment with cyclophosphamide in rat breast cancer.

BACKGROUND/AIM: It has been reported that continuous low-dose (metronomic) administration of cytotoxic drugs may be better tolerated and may have greater antitumor effects than a single high-dose chemotherapy. The aim of this study was to examine the efficacy and cardiotoxicity of metronomic administration of two of the most commonly used anticancer agents, cyclophosphamide (CPA) and doxorubicin (DOX), on an experimental breast cancer of rats. MATERIALS AND METHODS: Breast tumors were induced in Fisher 344 female rats by implanting Mat B III cells. Rats with tumors were randomized into three groups and were treated with a total dose of 160 mg/kg CPA and a total dose of 12 mg/kg DOX, administered twice per week for four weeks. Control rats were injected with saline according to the same schedule. Echocardiography was performed before the start of treatment and before sacrifice, which took place two weeks after the last injection, when plasma troponin was also measured. RESULTS: The metronomic CPA eradicated the tumors and preserved body weight and echocardiographic parameters. The metronomic DOX slowed tumor growth, but was not able to prevent DOX-induced cardiotoxicity. CONCLUSION: These results suggest that the success of a metronomic chemotherapy in terms of both efficacy and toxicity depends on the target, the class and the route of administration of the anticancer agent.

Tamoxifen and raloxifene suppress the proliferation of estrogen receptor-negative cells through inhibition of glutamine uptake.

PURPOSE: Modulation of estrogen receptor (ER) plays a central role in selective estrogen receptor modulators (SERMs) molecular mechanism of action, although studies have indicated that additional, non-ER-mediated mechanisms exist. It has been suggested that the induction of oxidative stress by SERM could be one of the non-ER-mediated mechanisms held responsible for their pro-apoptotic role in ER-negative cells. Tumor cells are known for their high requirement of glutamine (Gln) that serves multiple functions within the cells, including nutritional and energy source, as well as one of the precursors for the synthesis of natural antioxidant glutathione (GSH). We hypothesized that one of the mechanisms responsible for ER-independent anti-neoplastic properties of SERMs and also for their adverse side effects could be dependent on the inhibition of Gln uptake. METHODS: Human ER-negative MDA-MB231 breast cancer cells were treated with different doses of Tam and Ral. Gln uptake was monitored by using [(3)H]Gln assay. The effect of Tam and Ral on Gln transporter ASCT2 expression, glutathione (GSH) levels and cellular proliferation was determined. RESULTS: Tam and Ral inhibited Gln uptake in a dose-dependent manner through inhibition of ASCT2 Gln transporter. This effect of the anti-estrogens was associated with inhibition of GSH production and apoptosis. Treatment of cells with N-acetyl L-cysteine and 17 beta-estradiol 2 reversed the effects of Ral and Tam. CONCLUSIONS: Our results indicate that one of the mechanisms of action (and possibly some of the side effects) of TAM and RAL is associated with inhibition of cellular Gln uptake, oxidative stress and induction of apoptosis.

Glutamine regulation of doxorubicin accumulation in hearts versus tumors in experimental rats.

PURPOSE: Doxorubicin (DOX), an effective antineoplastic agent is known for its cardiotoxicity attributed mainly to free radical formation. Preliminary data indicated that oral glutamine (GLN) reduced cardiac oxidative damages in experimental rats treated with DOX. This study investigated the effect of GLN on DOX accumulation in tumors and normal tissues, troponin plasma concentration and functional alternations associated with DOX-induced myocardial damage. METHODS: Female Fisher344 rats (n = 40) with implanted MatBIII mammary tumors were randomized to receive oral GLN (1 g/kg/day) (n = 20) or to serve as controls (n = 20) and were treated with a single i.p. injection of 12 mg/kg DOX. Ten normal rats (n = 10) without treatment served as naive controls. Cardiac physiologic alterations resulting from DOX treatment in GLN-supplemented and control rats were assessed by micro-ultrasound imaging at 3 and 10 days after DOX injection. Ten rats from each GLN-supplemented and control groups were killed at 3 and 10 days after DOX administration. At killing, hearts, livers, spleens, kidneys, tumors and sera were examined for DOX concentration by measuring DOX natural fluorescence. Hearts were examined for Von Willebrand factor (vWF) expression using immunohistochemistry. RESULTS: Glutamine supplementation resulted in a significant reduction of DOX concentration in the normal tissues, without a significant effect on tumor DOX concentration. GLN-supplemented rats had lower plasma cTnI levels and lower cardiac levels of vWF. DOX-induced alterations in the echocardiographic parameters were significantly reduced in the GLN-supplemented rats. CONCLUSIONS: These data indicate that GLN supplement is able to reduce DOX-induced cardiac damage and thus to enhance DOX therapeutic effectiveness.

Effect of glutamine on gut glutathione fractional release in the implanted tumor model.

Cancer and its treatments cause a marked depletion of glutamine (GLN). However, dietary GLN can restore this loss and improve the outcomes of the treatments. The reasons behind this need to be investigated. GLN is suggested to involve in glutathione (GSH) synthesis. Fast-growing tumors alter gut GLN metabolism, but the effect of tumor growth on gut GSH release remains unknown. We hypothesized that gut GSH release would decrease in the tumor-bearing host and this downregulation would be antagonized by supplemental GLN. Female Fisher-344 rats were randomized to the groups: GLN + TUMOR, Freamine (FA) + TUMOR, GLN + SHAM, and FA + SHAM. The rats were implanted with MTF-7 mammary tumors as tumor-bearing groups, whereas the rats were sham operated as control groups. The rats were pair fed chow, gavaged with 1 g/kg/day GLN or an isonitrogenous FA. Tumor growth, blood and gut mucosa GLN, glutamate, and/or GSH were measured. The gut extractions, defined as the difference of concentrations across the gut, were calculated. Supplemental GLN enhanced the gut GLN uptake and GSH release with tumor growth and significantly increased blood and gut mucosa GLN and/or GSH concentrations. Our results demonstrate the important antioxidant role of GLN and thus may have significant implications in nutritional immune modulation in cancer patients.

Conductive interstitial thermal therapy device for surgical margin ablation: in vivo verification of a theoretical model.

PURPOSE: To demonstrate the efficacy and predictability of a new conductive interstitial thermal therapy (CITT) device to ablate surgical margins. METHOD: The temperature distributions during thermal ablation of CITT were calculated with finite element modelling in a geometrical representation of perfused tissue. The depth of ablation was derived using the Arrhenius and the Sapareto and Dewey (S&D) models for the temperature range of 90 to 150 degrees C. The female pig animal model was used to test the validity of the mathematical model. Breast tissues were ablated to temperatures in the range of 79-170 degrees C, in vivo. Triphenyltetrazolium chloride viability stain was used to delineate viable tissue from ablated regions and the ablation depths were measured using digital imaging. RESULTS: The calculations suggest that the CITT can be used to ablate perfused tissues to a 10-15 mm width within 20 minutes. The measured and calculated depths of ablation were statistically equivalent (99% confidence intervals) within +/- 1mm at 170 degrees C. At lower temperatures the equivalence between the model and the observations was within +/- 2 mm. CONCLUSION: The CITT device can reliably and uniformly ablate a 10-15 mm wide region of soft tissue. Thus, it can be used to secure negative margins following the resection of a primary tumor, which could impede local recurrences in the treatment of local diseases such as early staged, non-metastatic, breast cancer.

Modulation of p53 and c-myc in DMBA-induced mammary tumors by oral glutamine.

Previous studies established that oral glutamine (GLN) reduced tumor development in implantable and 7,12-dimethylbenz(a)anthracene (DMBA)-induced breast cancer models. This finding was associated with a decrease in tumor glutathione (GSH) levels, while maintaining normal gut, blood, and breast GSH. Alterations in GSH levels contribute to the control of apoptotic and cell cycle-regulating signaling. The aim of this study was to examine the role of dietary GLN on activation of p53 and c-myc, which play critical roles in cancer development and sensitivity to radiation and chemotherapy. Mammary gland carcinomas were induced in rats by DMBA. The rats were gavaged daily with GLN or water (controls), starting 1 wk prior DMBA-application and throughout the duration of the experiment (11 wk after DMBA). Tumor DNA was examined for mutations in p53 exons 5 and 6. Protein and mRNA levels of p53, p21(WAF1/CIP1), PTEN, IGF-IR, mdm2, and c-myc in tumors of GLN-supplemented rats were compared with those of the control rats (received water). The sequencing of p53 showed that it was wild type. Increased phosphorylation of p53, as well as higher mRNA and protein levels of p21(WAF1/CIP1), PTEN, and mdm2, and lower levels of IGF-IR were detected in tumors of GLN-supplemented rats vs. controls. Both phosphorylated c-myc and c-myc mRNA levels were reduced by GLN. The up-regulation of tumor p53 signaling and down-regulation of c-myc, in addition to previously established inhibition of Akt signaling in DMBA-breast cancer model, suggest that dietary GLN could be a useful approach for increasing the effectiveness of cancer treatment.