TIME COURSE OF SKELETAL MUSCLE LOSS AND OXIDATIVE STRESS IN RATS WITH WALKER 256 SOLID TUMOR

Reactive oxygen species oxidize proteins and modulate the proteasomal system in muscle-wasting cancer cachexia. On day 5 (D5), day 10 (D10), and day 14 (D14) after tumor implantation, skeletal muscle was evaluated. Carbonylated proteins and thiobarbituric acid reactive substances were measured. Chemiluminescence was employed for lipid hydroperoxide estimation. Glutathione, superoxide dismutase, and total radical antioxidant capacity were evaluated. The proteasomal system was assessed by mRNA atrogin-1 expression. Increased muscle wasting, lipid hydroperoxide, and superoxide dismutase, and decreased glutathione levels and total radical antioxidant capacity, were found on D5 in accordance with increased mRNA atrogin-1 expression. All parameters were significantly modified in animals treated with alpha-tocopherol. The elevation in aldehylde levels and carbonylated proteins observed on D10 were reversed by cc-tocopherol treatment. Oxidative stress may trigger signal transduction of the proteasomal system and cause protein oxidation. These pathways may be associated with the mechanism of muscle wasting that occurs in cancer cachexia. Muscle Nerve 42: 950-958, 2010

In vitro effect of low intensity laser on the cytotoxicity produced by substances released by bleaching gel

This in vitro study aimed to analyze the effect of different parameters of phototherapy with low intensity laser on the viability of human dental pulp fibroblasts under the effect of substances released by bleaching gel. Cells were seeded into 96 wells plates (1 x 10³ cells/well) and placed in contact with culture medium conditioned by a 35 % hydrogen peroxide bleaching gel for 40 minutes, simulating the clinical condition of the in-office bleaching treatment. Cells cultured in ideal growth conditions served as positive control group (PC), and the cells grown in conditioned medium and non-irradiated served as negative control group (NC). Cells grown in conditioned medium were submitted to a single irradiation with a diode laser (40 mW, 0.04 cm²) emitting at visible red (660 nm; RL) or near infrared (780 nm; NIR) using punctual technique, in contact mode and energy densities of 4, 6 or 10 J/cm². The cell viability was analyzed through the MTT reduction assay immediately and 24 hours after the irradiation. The data was compared by ANOVA followed by the Tukey's test (p < 0.05). The cell viability increased significantly in 24 hours within each group. The PC presented cell viability significantly higher than NC in both experimental times. Only the NIR/10 J/cm² group presented cell viability similar to that of PC in 24 hours. The phototherapy with low intensity laser in defined parameters is able to compensate the cytotoxic effects of substances released by 35 % hydrogen peroxide bleaching gel.

Survival and quality of life of patients with oral and oropharyngeal cancer at 1-year follow-up of tumor resection

OBJECTIVE: This study aimed to assess the survival and life quality evolution of patients subjected to surgical excision of oral and oropharyngeal squamous cell carcinoma. MATERIAL AND METHODS: Forty-seven patients treated at a Brazilian healthcare unit specialized in head and neck surgery between 2006 and 2007 were enrolled in the study. The gathering of data comprised reviewing hospital files and applying the University of Washington Quality of Life (UW-QOL) questionnaire previously and 1 year after the surgery. Comparative analysis used Poisson regression to assess factors associated with survival and a paired t-test to compare preoperative and 1-year postoperative QOL ratings. RESULTS: 1 year after surgery, 7 patients were not found (dropout of the cohort); 15 had died and 25 fulfilled the UW-QOL again. The risk of death was associated with having regional metastasis previously to surgery (relative risk=2.18; 95% confidence interval=1.09-5.17) and tumor size T3 or T4 (RR=2.30; 95%CI=1.05-5.04). Survivors presented significantly (p<0.05) poorer overall and domain-specific ratings of quality of life. Chewing presented the largest reduction: from 74.0 before surgery to 34.0 one year later. Anxiety was the only domain whose average rating increased (from 36.0 to 70.7). CONCLUSIONS: The prospective assessment of survival and quality of life may contribute to anticipate interventions aimed at reducing the incidence of functional limitations in patients with oral and oropharyngeal cancer.

Effect of nitric oxide inhibitor and donor substances on the infammatory process caused by endodontic irrigants

Nitric oxide (NO) has been considered a key molecule in infammation. OBJECTIVE: The aim of this study was to evaluate the effect of treatment with L-NAME and sodium nitroprussiate, substances that inhibit and release NO, respectively, on tissue tolerance to endodontic irrigants. MATERIAL AND METHODS: The vital dye exudation method was used in a rat subcutaneous tissue model. Injections of 2% Evans blue were administered intravenously into the dorsal penial vein of 14 male rats (200-300 g). The NO inhibitor and donor substances were injected into the subcutaneous tissue in the dorsal region, forming two groups of animals: G1 was inoculated with L-NAME and G2 with sodium nitroprussiate. Both groups received injections of the test endodontic irrigants: acetic acid, 15% citric acid, 17% EDTA-T and saline (control). After 30 min, analysis of the extravasated dye was performed by light absorption spectrophotometry (620 nm). RESULTS: There was statistically signifcant difference (p<0.05) between groups 1 and 2 for all irrigants. L-NAME produced a less intense infammatory reaction and nitroprussiate intensifed this process. CONCLUSIONS: Independently of the administration of NO inhibitors and donors, EDTA-T produced the highest irritating potential in vital tissue among the tested irrigating solutions.

Impact of adiposity on immunological parameters

Studies evaluating immune function in obese humans and experimental animals indicate that the excess adiposity is associated with impaired in immune responses. Obesity is related to a higher rate of infections and to some types of cancer. Nutritional, metabolic and endocrine factors are implicated in the immunological changes. The adipose tissue directly produces substances with various functions related to immune system. Furthermore, some investigations suggest that certain types of weight reduction strategies can alter the immune function. Nevertheless, long-term studies should be carried out to address whether these changes positively affects the ability of these obese individuals to control infections and tumor development.

Effect of cyhalothrin on Ehrlich tumor growth and macrophage activity in mice

Cyhalothrin, a pyrethroid insecticide, induces stress-like symptoms, increases c-fos immunoreactivity in the paraventricular nucleus of the hypothalamus, and decreases innate immune responses in laboratory animals. Macrophages are key elements in cellular immune responses and operate at the tumor-host interface. This study investigated the relationship among cyhalothrin effects on Ehrlich tumor growth, serum corticosterone levels and peritoneal macrophage activity in mice. Three experiments were done with 10 experimental (single gavage administration of 3.0 mg/kg cyhalothrin daily for 7 days) and 10 control (single gavage administration of 1.0 mL/kg vehicle of cyhalothrin preparation daily for 7 days) isogenic BALB/c mice in each experiment. Cyhalothrin i) increased Ehrlich ascitic tumor growth after ip administration of 5.0 x 106 tumor cells, i.e., ascitic fluid volume (control = 1.97 ± 0.39 mL and experimental = 2.71 ± 0.92 mL; P < 0.05), concentration of tumor cells/mL in the ascitic fluid (control = 111.95 ± 16.73 x 106 and experimental = 144.60 ± 33.18 x 106; P < 0.05), and total number of tumor cells in the ascitic fluid (control = 226.91 ± 43.22 x 106 and experimental = 349.40 ± 106.38 x 106; P < 0.05); ii) increased serum corticosterone levels (control = 200.0 ± 48.3 ng/mL and experimental = 420.0 ± 75.5 ng/mL; P < 0.05), and iii) decreased the intensity of macrophage phagocytosis (control = 132.3 ± 19.7 and experimental = 116.2 ± 4.6; P < 0.05) and oxidative burst (control = 173.7 ± 40.8 and experimental= 99.58 ± 41.7; P < 0.05) in vitro in the presence of Staphylococcus aureus. These data provide evidence that cyhalothrin simultaneously alters host resistance to Ehrlich tumor growth, hypothalamic-pituitary-adrenocortical (HPA) axis function, and peritoneal macrophage activity. The results are discussed in terms of data suggesting a link between stress, HPA axis activation and resistance to tumor growth.

Gene expression down-regulation in CD90(+) prostate tumor-associated stromal cells involves potential organ-specific genes

Background: The prostate stroma is a key mediator of epithelial differentiation and development, and potentially plays a role in the initiation and progression of prostate cancer. The tumor-associated stroma is marked by increased expression of CD90/THYI. Isolation and characterization of these stromal cells could provide valuable insight into the biology of the tumor microenvironment. Methods: Prostate CD90(+) stromal fibromuscular cells from tumor specimens were isolated by cell-sorting and analyzed by DNA microarray. Dataset analysis was used to compare gene expression between histologically normal and tumor-associated stromal cells. For comparison, stromal cells were also isolated and analyzed from the urinary bladder. Results: The tumor-associated stromal cells were found to have decreased expression of genes involved in smooth muscle differentiation, and those detected in prostate but not bladder. Other differential expression between the stromal cell types included that of the CXC-chemokine genes. Conclusion: CD90(+) prostate tumor-associated stromal cells differed from their normal counterpart in expression of multiple genes, some of which are potentially involved in organ development.

A perivascular epithelioid cell tumor of the stomach: An unsuspected diagnosis

Perivascular epithelioid cell tumor (PEComa) is a rare mesenchymal neoplasia and currently well recognized as a distinct entity with characteristic morphological, immunohistochemical and molecular findings. We report a case of PEComa arising in the antrum of a 71-year-old female with melena. The tumor, located predominantly in the submucosa as a well delimited nodule, measured 3.0 cm in diameter and was completely resected, with no evidence of the disease elsewhere. Histologically, it was composed predominantly of eosinophilic epithelioid cells arranged in small nests commonly related to variably sized vessels, with abundant extracellular material, moderate nuclear variation and discrete mitotic activity. No necrosis, angiolymphatic invasion or perineural infiltration was seen. Tumor cells were uniformly positive for vimentin, smooth muscle actin, desmin and melan A. Although unusual, PEComa should be considered in the differential diagnosis of gastric neoplasia with characteristic epithelioid and oncocytic features and prominent vasculature: (C) 2010 Baishideng. All rights reserved.

Evaluation of the Incidence of Bladder Perforation After Transurethral Bladder Tumor Resection in a Residency Setting

Purpose: To evaluate prospectively the actual bladder perforation incidence during transurethral resection of bladder tumor (TURB) performed by residents and to identify possible predisposing factors to such condition. Patients and Methods: Thirty-four patients with bladder tumor were submitted to TURB in our academic institution in April 2006, and were prospectively studied. Procedures were all done by senior residents under an attending direct supervision. All patients had a cystograms performed after the procedure by the injection of 400 mL of saline-diluted contrast solution with low-pressure infusion through the Foley catheter. The cystograms were evaluated blindly by a single radiologist. All patients were examined by cystoscopy and/or CT every 3 months for the first 2 years postoperatively. Results: The cystogram showed contrast leaking compatible with bladder perforation in 17 (50%) cases. None of the perforations were recognized intraoperatively by the surgeon. All perforations were extraperitoneal and managed conservatively. There was no significant correlation between the incidence of bladder perforation and the patient age (p = 0.508), the tumor stage (p = 0.998), the tumor grade (p = 0.833), the number of lesions (p = 0.394), and the tumor size (p = 0.651). The only factor that had impact on the development of bladder perforation was tumor localization at the bottom of the bladder (p = 0.035; OR, 6750; 95% CI, 1.14, 39.8). Conclusion: Asymptomatic perforations of the bladder wall occur very frequently after a TURB procedure performed by residents in training and, most of the time, are not noticed by the surgeon. Localization of the tumor at bladder dome was the only factor that negatively influenced perforation rates.

Anti-tumor therapy with macroencapsulated endostatin producer cells

Background: Theracyte is a polytetrafluoroethylene membrane macroencapsulation system designed to induce neovascularization at the tissue interface, protecting the cells from host's immune rejection, thereby circumventing the problem of limited half-life and variation in circulating levels. Endostatin is a potent inhibitor of angiogenesis and tumor growth. Continuous delivery of endostatin improves the efficacy and potency of the antitumoral therapy. The purpose of this study was to determine whether recombinant fibroblasts expressing endostatin encapsulated in Theracyte immunoisolation devices can be used for delivery of this therapeutic protein for treatment of mice bearing B16F10 melanoma and Ehrlich tumors. Results: Mice were inoculated subcutaneously with melanoma (B16F10 cells) or Ehrlich tumor cells at the foot pads. Treatment began when tumor thickness had reached 0.5 mm, by subcutaneous implantation of 10(7) recombinant encapsulated or non-encapsulated endostatin producer cells. Similar melanoma growth inhibition was obtained for mice treated with encapsulated or non-encapsulated endostatin-expressing cells. The treatment of mice bearing melanoma tumor with encapsulated endostatin-expressing cells was decreased by 50.0%, whereas a decrease of 56.7% in tumor thickness was obtained for mice treated with non-encapsulated cells. Treatment of Ehrlich tumor-bearing mice with non-encapsulated endostatin-expressing cells reduced tumor thickness by 52.4%, whereas lower tumor growth inhibition was obtained for mice treated with encapsulated endostatin-expressing cells: 24.2%. Encapsulated endostatin-secreting fibroblasts failed to survive until the end of the treatment. However, endostatin release from the devices to the surrounding tissues was confirmed by immunostaining. Decrease in vascular structures, functional vessels and extension of the vascular area were observed in melanoma microenvironments. Conclusions: This study indicates that immunoisolation devices containing endostatin-expressing cells are effective for the inhibition of the growth of melanoma and Ehrlich tumors. Macroencapsulation of engineered cells is therefore a reliable platform for the refinement of innovative therapeutic strategies against tumors.

Evaluation of Dendritic Cell-Tumor Cell Hybrid Vaccine Storage

Clinical trials using dendritic cells (DCs) to treat cancer patients have generated promising results in recent years. However, even simple aspects of this therapy are still not well understood, including the storage and distribution of manufactured vaccines. These processes are essential and must be elucidated in order to reduce costs. We evaluated the effects of different storage conditions on vaccine functionality using mixed lymphocyte reaction (MLR). Vaccine storage at 4 degrees C for up to 72 h had no significant effect on vaccine activity. Shipping to distant places is possible, if vaccines are kept at 4 degrees C and used up to 3 days after manufacture date.

Interleukin-10 production by tumor infiltrating macrophages plays a role in Human Papillomavirus 16 tumor growth

Background: Human Papillomavirus, HPV, is the main etiological factor for cervical cancer. Different studies show that in women infected with HPV there is a positive correlation between lesion grade and number of infiltrating macrophages, as well as with IL-10 higher expression. Using a HPV16 associated tumor model in mice, TC-1, our laboratory has demonstrated that tumor infiltrating macrophages are M2-like, induce T cell regulatory phenotype and play an important role in tumor growth. M2 macrophages secrete several cytokines, among them IL-10, which has been shown to play a role in T cell suppression by tumor macrophages in other tumor models. In this work, we sought to establish if IL-10 is part of the mechanism by which HPV tumor associated macrophages induce T cell regulatory phenotype, inhibiting anti-tumor activity and facilitating tumor growth. Results: TC-1 tumor cells do not express or respond to IL-10, but recruit leukocytes which, within the tumor environment, produce this cytokine. Using IL-10 deficient mice or blocking IL-10 signaling with neutralizing antibodies, we observed a significant reduction in tumor growth, an increase in tumor infiltration by HPV16 E7 specific CD8 lymphocytes, including a population positive for Granzyme B and Perforin expression, and a decrease in the percentage of HPV specific regulatory T cells in the lymph nodes. Conclusions: Our data shows that in the HPV16 TC-1 tumor mouse model, IL-10 produced by tumor macrophages induce regulatory phenotype on T cells, an immune escape mechanism that facilitates tumor growth. Our results point to a possible mechanism behind the epidemiologic data that correlates higher IL-10 expression with risk of cervical cancer development in HPV infected women.

Tetraploidization in Wilms tumor in an infant

Genetic instability is frequent in human cancer. Unscheduled tetraploidization can trigger cell transformation and tumorigenesis. We made a cytogenetic analysis by Giemsa-trypsin banding of a stage I, biphasic Wilms tumor diagnosed in a 10-month-old male. An evident karyotypic heterogeneity was found. Four different subclones of tumor cells were observed, with DNA content varying from diploid to near-tetraploid complements. The genetic events involved in the acquisition of aneuploidy in Wilms tumor remain unclear. We hypothesize that initial tetraploidization caused aberrant cell division, leading to abnormal chromosomal segregation, cell transformation and tumorigenesis.

Tumor slices as a model to evaluate doxorubicin in vitro treatment and expression of trios of genes PRSS11, MTSS1, CLPTM1 and PRSS11, MTSS1, SMYD2 in canine mammary gland cancer

Background: In women with breast cancer submitted to neoadjuvant chemotherapy based in doxorubicin, tumor expression of groups of three genes (PRSS11, MTSS1, CLPTM1 and PRSS11, MTSS1, SMYD2) have classified them as responsive or resistant. We have investigated whether expression of these trios of genes could predict mammary carcinoma response in dogs and whether tumor slices, which maintain epithelial-mesenchymal interactions, could be used to evaluate drug response in vitro. Methods: Tumors from 38 dogs were sliced and cultured with or without doxorubicin 1 mu M for 24 h. Tumor cells were counted by two observers to establish a percentage variation in cell number, between slices. Based on these results, a reduction in cell number between treated and control samples >= 21.7%, arbitrarily classified samples, as drug responsive. Tumor expression of PRSS11, MTSS1, CLPTM1 and SMYD2, was evaluated by real time PCR. Relative expression results were then transformed to their natural logarithm values, which were spatially disposed according to the expression of trios of genes, comprising PRSS11, MTSS1, CLPTM1 and PRSS11, MTSS1, SMYD2. Fisher linear discrimination test was used to generate a separation plane between responsive and non-responsive tumors. Results: Culture of tumor slices for 24 h was feasible. Nine samples were considered responsive and 29 non-responsive to doxorubicin, considering the pre-established cut-off value of cell number reduction = 21.7%, between doxorubicin treated and control samples. Relative gene expression was evaluated and tumor samples were then spatially distributed according to the expression of the trios of genes: PRSS11, MTSS1, CLPTM1 and PRSS11, MTSS1, SMYD2. A separation plane was generated. However, no clear separation between responsive and non-responsive samples could be observed. Conclusion: Three-dimensional distribution of samples according to the expression of the trios of genes PRSS11, MTSS1, CLPTM1 and PRSS11, MTSS1, SMYD2 could not predict doxorubicin in vitro responsiveness. Short term culture of mammary gland cancer slices may be an interesting model to evaluate chemotherapy activity.

Activation of endogenous p53 by combined p19Arf gene transfer and nutlin-3 drug treatment modalities in the murine cell lines B16 and C6

Background: Reactivation of p53 by either gene transfer or pharmacologic approaches may compensate for loss of p19Arf or excess mdm2 expression, common events in melanoma and glioma. In our previous work, we constructed the pCLPG retroviral vector where transgene expression is controlled by p53 through a p53-responsive promoter. The use of this vector to introduce p19Arf into tumor cells that harbor p53wt should yield viral expression of p19Arf which, in turn, would activate the endogenous p53 and result in enhanced vector expression and tumor suppression. Since nutlin-3 can activate p53 by blocking its interaction with mdm2, we explored the possibility that the combination of p19Arf gene transfer and nutlin-3 drug treatment may provide an additive benefit in stimulating p53 function. Methods: B16 (mouse melanoma) and C6 (rat glioma) cell lines, which harbor p53wt, were transduced with pCLPGp19 and these were additionally treated with nutlin-3 or the DNA damaging agent, doxorubicin. Viral expression was confirmed by Western, Northern and immunofluorescence assays. p53 function was assessed by reporter gene activity provided by a p53-responsive construct. Alterations in proliferation and viability were measured by colony formation, growth curve, cell cycle and MTT assays. In an animal model, B16 cells were treated with the pCLPGp19 virus and/or drugs before subcutaneous injection in C57BL/6 mice, observation of tumor progression and histopathologic analyses. Results: Here we show that the functional activation of endogenous p53wt in B16 was particularly challenging, but accomplished when combined gene transfer and drug treatments were applied, resulting in increased transactivation by p53, marked cell cycle alteration and reduced viability in culture. In an animal model, B16 cells treated with both p19Arf and nutlin-3 yielded increased necrosis and decreased BrdU marking. In comparison, C6 cells were quite susceptible to either treatment, yet p53 was further activated by the combination of p19Arf and nutlin-3. Conclusions: To the best of our knowledge, this is the first study to apply both p19Arf and nutlin-3 for the stimulation of p53 activity. These results support the notion that a p53 responsive vector may prove to be an interesting gene transfer tool, especially when combined with p53- activating agents, for the treatment of tumors that retain wild-type p53.