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Asian Pacific Journal of Cancer... Oct 2020To analyze the effect of sirolimus and sunitinib in blocking the tumor growth and to evaluate the expressions of estrogen receptor (ER), progesterone receptor (PgR), and...
OBJECTIVE
To analyze the effect of sirolimus and sunitinib in blocking the tumor growth and to evaluate the expressions of estrogen receptor (ER), progesterone receptor (PgR), and human epidermal growth factor receptor-2 (HER2/neu) after treated with sirolimus and sunitinib.
METHODS
Thirty-two female Sprague Dawley rats at age 21-days old were administered intraperitoneally with N-Methyl-N-Nitroso Urea (NMU), dosed at 70mg/kg body weight. The rats were divided into 4 groups; Group 1 (Control, n=8), Group 2 (Sirolimus, n=8), Group 3 (Sunitinib, n=8) and Group 4 (Sirolimus+Sunitinib, n=8), being treated twice when the tumor reached the size of 14.5±0.5 mm and subsequently sacrificed after 5 days. The protein expressions of ER, PgR and HER2/neu of the tumor tissues were evaluated by using immunohistochemistry analysis.
RESULTS
Treatment with sirolimus alone lowered expressions of ER and PgR of breast cancer and reduced tumor size. There was no significant difference of ER and PgR expressions between control and sunitinib treated tumor. Sunitinib treated tumors reduce in diameter after the first treatment, however the diameter increases after the second treatment. Histologically, sunitinib treated tumor did not show any aggressive invasive carcinoma of no special type (NST) histological subtypes. In addition, all NMU-induced tumors are HER2/neu-negative scoring.
CONCLUSION
Sirolimus is neither synergistic nor additive with sunitinib for breast cancer treatment.
.Topics: Alkylating Agents; Animals; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Cell Proliferation; Female; Mammary Neoplasms, Experimental; Methylnitrosourea; Rats; Rats, Sprague-Dawley; Receptor, ErbB-2; Receptors, Estrogen; Receptors, Progesterone; Sirolimus; Sunitinib
PubMed: 33112549
DOI: 10.31557/APJCP.2020.21.10.2919 -
Scientific Reports Dec 2021Retinal prosthesis is regarded as the treatment for vision restoration in the blind with retinal degeneration (RD) due to the loss of photoreceptors. A strategy for...
Retinal prosthesis is regarded as the treatment for vision restoration in the blind with retinal degeneration (RD) due to the loss of photoreceptors. A strategy for retinal prosthesis is to electrically activate surviving neurons. The retina's response to electrical stimulation in a larger RD model has not been studied yet. Therefore, in this study, we investigated electrically evoked retinal responses in a previously validated N-methyl-N-nitrosourea (MNU)-induced porcine RD model. Electrically evoked responses were evaluated based on the number of retinal ganglion cell (RGC) spikes via multichannel recordings. Stimulation pulses were applied to degenerative and wild-type retinas with pulse modulation. Compared to wild-type retinas, degenerative retinas showed higher threshold values of pulse amplitude and pulse duration. The rate of increase in the number of RGC spikes relative to stimulus intensity was lower in degenerative retinas than in normal retinas. In severely degenerated retinas, few RGCs showed electrically evoked spikes. Our results suggest that the degenerative porcine retina requires a higher charge than the normal porcine retina. In the early stage of RD, it is easier to induce RGC spikes through electrical stimulation using retinal prosthesis; however, when the degeneration is severe, there may be difficulty recovering patient vision.
Topics: Animals; Disease Models, Animal; Electric Stimulation; Evoked Potentials, Visual; Female; Methylnitrosourea; Retinal Degeneration; Retinal Ganglion Cells; Swine; Swine, Miniature
PubMed: 34921172
DOI: 10.1038/s41598-021-03439-w -
The Journal of Biological Chemistry Jan 2020The mismatch repair (MMR) complex is composed of MutSα (MSH2-MSH6) and MutLα (MLH1-PMS2) and specifically recognizes mismatched bases during DNA replication....
The mismatch repair (MMR) complex is composed of MutSα (MSH2-MSH6) and MutLα (MLH1-PMS2) and specifically recognizes mismatched bases during DNA replication. -Methylguanine is produced by treatment with alkylating agents, such as -methyl--nitrosourea (MNU), and during DNA replication forms a DNA mismatch ( an -methylguanine/thymine pair) and induces a G/C to A/T transition mutation. To prevent this outcome, cells carrying this DNA mismatch are eliminated by MMR-dependent apoptosis, but the underlying molecular mechanism is unclear. In this study, we provide evidence that the chromatin-regulatory and ATP-dependent nucleosome-remodeling protein SMARCAD1 is involved in the induction of MMR-dependent apoptosis in human cells. Unlike control cells, -knockout cells (ΔSMARCAD1) were MNU-resistant, and the appearance of a sub-G population and caspase-9 activation were significantly suppressed in the ΔSMARCAD1 cells. Furthermore, the MNU-induced mutation frequencies were increased in these cells. Immunoprecipitation analyses revealed that the recruitment of MutLα to chromatin-bound MutSα, observed in SMARCAD1-proficient cells, is suppressed in ΔSMARCAD1 cells. Of note, the effect of SMARCAD1 on the recruitment of MutLα exclusively depended on the ATPase activity of the protein. On the basis of these findings, we propose that SMARCAD1 induces apoptosis via its chromatin-remodeling activity, which helps recruit MutLα to MutSα on damaged chromatin.
Topics: Apoptosis; Cell Line, Tumor; Chromatin; DNA Damage; DNA Helicases; DNA Mismatch Repair; Gene Knockout Techniques; Humans; Methylnitrosourea; Models, Biological; MutL Proteins; MutS Homolog 2 Protein; Mutation Rate; Signal Transduction
PubMed: 31843968
DOI: 10.1074/jbc.RA119.008854 -
Metabolites Jul 2019Metabolomics is an effective approach to characterize the metabotype which can reflect the influence of genetics, physiological status, and environmental factors such as...
Metabolomics is an effective approach to characterize the metabotype which can reflect the influence of genetics, physiological status, and environmental factors such as drug intakes, diet. Diet may change the chemopreventive efficacy of given agents due to the altered physiological status of the subject. Here, metabolomics response to a chemopreventive agent targretin or tamoxifen, in rats with methylnitrosourea-induced tumors on a standard diet (4% fat, CD) or a high fat diet (21% fat, HFD) was evaluated, and found that (1) the metabolome was substantially affected by diet and/or drug treatment; (2) multiple metabolites were identified as potential pharmacodynamic biomarkers related to targretin or tamoxifen regardless of diet and time; and (3) the primary bile acid pathway was significantly affected by targretin treatment in rats on both diets, and the lysolipid pathway was significantly affected by tamoxifen treatment in rats on the high fat diet.
PubMed: 31336604
DOI: 10.3390/metabo9070149 -
Chemical Research in Toxicology Feb 2020DNA methylating agents are abundant in the environment and are sometimes used in cancer chemotherapy. They react with DNA to form methyl-DNA adducts and byproduct...
DNA methylating agents are abundant in the environment and are sometimes used in cancer chemotherapy. They react with DNA to form methyl-DNA adducts and byproduct lesions that can be both toxic and mutagenic. Foremost among the mutagenic lesions is -methylguanine (m6G), which base pairs with thymine during replication to cause GC → AT mutations. The delta C57BL/6J mouse strain of Nohmi et al. ( , , 465-70) reliably produces mutational spectra of many DNA damaging agents. In this work, mouse embryo fibroblasts (MEFs) were made from delta C57BL/6J mice and evaluated as a screening tool to determine the qualitative and quantitative features of mutagenesis by -methyl--nitrosourea (MNU), a direct-acting DNA alkylator that serves as a model for environmental -nitrosamines, such as -nitrosodimethylamine and therapeutic agents such as Temozolomide. The DNA repair protein MGMT (-methylguanine DNA methyltransferase) protects against environmental mutagenesis by DNA methylating agents and, by removing m6G, limits the therapeutic potential of Temozolomide in cancer therapy. The delta MEFs were treated with MNU to establish dose-dependent toxicity. In parallel, MNU mutagenicity was determined in the presence and absence of the MGMT inhibitor AA-CW236 (4-(2-(5-(chloromethyl)-4-(4-(trifluoromethoxy)phenyl)-1H-1,2,3-triazol-1-yl)ethyl)-3,5-dimethylisoxazole). With and without the inhibitor, the principal mutagenic event of MNU was GC → AT, but more mutations were observed when the inhibitor was present. Evidence that the mutagenic lesion was m6G was based on mass spectral data collected using -methyl--guanine as an internal standard; m6G levels were higher in AA-CW236 treated MEFs by an amount proportional to the higher mutation frequency seen in the same cells. This work establishes delta MEFs as a versatile tool for probing mutagenesis by environmental and therapeutic agents and as a cell culture model in which chemical genetics can be used to determine the impact of DNA repair on biological responses to DNA damaging agents.
Topics: Alkylating Agents; Animals; DNA Modification Methylases; DNA Repair Enzymes; Enzyme Inhibitors; Fibroblasts; Methylnitrosourea; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mutagenesis; Tumor Suppressor Proteins
PubMed: 31841318
DOI: 10.1021/acs.chemrestox.9b00444 -
BMC Molecular and Cell Biology Jun 2021In the present study, fatty acid synthesis is targeted to combat mammary gland carcinoma by activating prolyl hydroxylase-2 with Voacamine alone and in combination with...
BACKGROUND
In the present study, fatty acid synthesis is targeted to combat mammary gland carcinoma by activating prolyl hydroxylase-2 with Voacamine alone and in combination with Tamoxifen. It was hypothesized that the activation of prolyl hydroxylase-2 would inhibit the hypoxia-induced fatty acid synthesis and mammary gland carcinoma. Mammary gland carcinoma was induced with a single dose administration of N-methyl-N-nitrosourea (50 mg/kg,i.p.) and treatment with Voacamine and Tamoxifen 15 days after carcinogen administration.
RESULTS
At the end of the study, hemodynamic profiling of animals was recorded to assess the cardiotoxic potential of the drug. Blood serum was separated and subjected to nuclear magnetic resonance spectroscopy. Carmine staining and histopathology of mammary gland tissue were performed to evaluate the anti-angiogenic potential of the drug. The antioxidant potential of the drug was measured with antioxidant markers. Western blotting was performed to study the effect of the drug at the molecular level.
CONCLUSION
Results of the study have shown that Voacamine treatment stopped further decrease in body weight of experimental animals. The hemodynamic study evidenced that Voacamine at a low dose is safe in cardiac patients. Microscopic evaluation of mammary gland tissue documented the anti-angiogenic potential of Voacamine and Tamoxifen therapy. Perturbed serum metabolites were also restored to normal along with antioxidant markers. Immunoblotting of mammary gland tissue also depicted restoration of proteins of the hypoxic and fatty acid pathway. Conclusively, Voacamine and its combination with Tamoxifen activated prolyl hydroxylase-2 to combat mammary gland carcinoma.
Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Antioxidants; Body Weight; Carcinoma; Computer Simulation; Electrocardiography; Fatty Acids; Female; Heart Rate; Hypoxia-Inducible Factor-Proline Dioxygenases; Ibogaine; Mammary Glands, Animal; Mammary Neoplasms, Experimental; Metabolome; Methylnitrosourea; Neovascularization, Pathologic; Rats, Wistar; Tamoxifen; Rats
PubMed: 34090331
DOI: 10.1186/s12860-021-00371-9 -
Graefe's Archive For Clinical and... May 2015To identify programmed cell death (PCD) pathways involved in N-methyl-N-nitrosourea (MNU)-induced photoreceptor (PR) degeneration.
PURPOSE
To identify programmed cell death (PCD) pathways involved in N-methyl-N-nitrosourea (MNU)-induced photoreceptor (PR) degeneration.
METHODS
Adult C57BL/6 mice received a single MNU i.p. injection (60 mg/kg bodyweight), and were observed over a period of 7 days. Degeneration was visualized by H&E overview staining and electron microscopy. PR cell death was measured by quantifying TUNEL-positive cells in the outer nuclear layer (ONL). Activity measurements of key PCD enzymes (calpain, caspases) were used to identify the involved cell death pathways. Furthermore, the expression level of C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (GRP78), key players in endoplasmic reticulum (ER) stress-induced apoptosis, was analyzed using quantitative real-time PCR.
RESULTS
A decrease in ONL thickness and the appearance of apoptotic PR nuclei could be detected beginning 3 days post-injection (PI). This was accompanied by an increase of TUNEL-positive cells. Significant upregulation of activated caspases (3, 9, 12) was found at different time periods after MNU injection. Additionally, several other players of nonconventional PCD pathways were also upregulated. Consequently, calpain activity increased in the ONL, with a maximum on day 7 PI and an upregulation of CHOP and GRP78 expression beginning on day 1 PI was found.
CONCLUSIONS
The data indicate that regular apoptosis is the major cause of MNU-induced PR cell death. However, alternative PCD pathways, including ER stress and calpain activation, are also involved. Knowledge about the mechanisms involved in this mouse model of PR degeneration could facilitate the design of putative combinatory therapeutic approaches.
Topics: Alkylating Agents; Animals; Apoptosis; Calpain; Caspases; Disease Models, Animal; Endoplasmic Reticulum Chaperone BiP; Heat-Shock Proteins; Humans; In Situ Nick-End Labeling; Injections, Intraperitoneal; Methylnitrosourea; Mice; Mice, Inbred C57BL; Photoreceptor Cells, Vertebrate; RNA, Messenger; Real-Time Polymerase Chain Reaction; Retinal Degeneration; Transcription Factor CHOP
PubMed: 25875043
DOI: 10.1007/s00417-014-2906-x -
Nature Jan 2015Next-generation sequencing of human tumours has refined our understanding of the mutational processes operative in cancer initiation and progression, yet major questions...
Next-generation sequencing of human tumours has refined our understanding of the mutational processes operative in cancer initiation and progression, yet major questions remain regarding the factors that induce driver mutations and the processes that shape mutation selection during tumorigenesis. Here we performed whole-exome sequencing on adenomas from three mouse models of non-small-cell lung cancer, which were induced either by exposure to carcinogens (methyl-nitrosourea (MNU) and urethane) or by genetic activation of Kras (Kras(LA2)). Although the MNU-induced tumours carried exactly the same initiating mutation in Kras as seen in the Kras(LA2) model (G12D), MNU tumours had an average of 192 non-synonymous, somatic single-nucleotide variants, compared with only six in tumours from the Kras(LA2) model. By contrast, the Kras(LA2) tumours exhibited a significantly higher level of aneuploidy and copy number alterations compared with the carcinogen-induced tumours, suggesting that carcinogen-induced and genetically engineered models lead to tumour development through different routes. The wild-type allele of Kras has been shown to act as a tumour suppressor in mouse models of non-small-cell lung cancer. We demonstrate that urethane-induced tumours from wild-type mice carry mostly (94%) Kras Q61R mutations, whereas those from Kras heterozygous animals carry mostly (92%) Kras Q61L mutations, indicating a major role for germline Kras status in mutation selection during initiation. The exome-wide mutation spectra in carcinogen-induced tumours overwhelmingly display signatures of the initiating carcinogen, while adenocarcinomas acquire additional C > T mutations at CpG sites. These data provide a basis for understanding results from human tumour genome sequencing, which has identified two broad categories of tumours based on the relative frequency of single-nucleotide variations and copy number alterations, and underline the importance of carcinogen models for understanding the complex mutation spectra seen in human cancers.
Topics: Adenocarcinoma; Animals; Carcinogens; Carcinoma, Non-Small-Cell Lung; Cell Transformation, Neoplastic; DNA Copy Number Variations; Disease Progression; Female; Genes, ras; Genomic Instability; Germ-Line Mutation; Humans; Lung Neoplasms; Male; Methylnitrosourea; Mice; Models, Genetic; Mutation; Oncogene Protein p21(ras); Point Mutation; Proto-Oncogene Proteins p21(ras); Urethane
PubMed: 25363767
DOI: 10.1038/nature13898 -
Drug Delivery Dec 2021Retinal degeneration (RD) refers to a group of blinding retinopathies leading to the progressive photoreceptor demise and vision loss. Treatments against this...
Retinal degeneration (RD) refers to a group of blinding retinopathies leading to the progressive photoreceptor demise and vision loss. Treatments against this debilitating disease are urgently needed. Intraocular delivery of exosomes represents an innovative therapeutic strategy against RD. In this study, we aimed to determine whether the subretinal delivery of RPE-derived exosomes (RPE-Exos) can prevent the photoreceptor death in RD. RD was induced in C57BL6 mice by MNU administration. These MNU administered mice received a single subretinal injection of RPE-Exos. Two weeks later, the RPE-Exos induced effects were evaluated via functional, morphological, and behavior examinations. Subretinal delivery of RPE-Exos efficiently ameliorates the visual function impairments, and alleviated the structural damages in the retina of MNU administered mice. Moreover, RPE-Exos exert beneficial effects on the electrical response of the inner retinal circuits. Treatment with RPE-Exos suppressed the expression levels of inflammatory factors, and mitigated the oxidative damage, indicating that subretinal delivery of RPE-Exos constructed a cytoprotective microenvironment in the retina of MNU administered mice. Our data suggest that RPE-Exos have therapeutic effects against the visual impairments and photoreceptor death. These findings will enrich our knowledge of RPE-Exos, and highlight the discovery of a promising medication for RD.
Topics: Alkylating Agents; Animals; Apoptosis; Biological Products; Calpain; Caspase 3; Disease Models, Animal; Electroretinography; Exosomes; Inflammation; Injections, Intraocular; Interleukin-1beta; Interleukin-6; Malondialdehyde; Methylnitrosourea; Mice; Oxidative Stress; Photoreceptor Cells, Vertebrate; Proto-Oncogene Proteins c-bcl-2; Retina; Retinal Degeneration; Retinal Pigment Epithelium; Tomography, Optical Coherence; Tumor Necrosis Factor-alpha; Vision, Ocular; bcl-2-Associated X Protein
PubMed: 33501868
DOI: 10.1080/10717544.2020.1870584 -
Cancer Immunology Research Jul 2017Intravesical bacillus Calmette-Guérin (BCG) immunotherapy is the standard of care in treating non-muscle-invasive bladder cancer, yet its mechanism of action remains...
Intravesical bacillus Calmette-Guérin (BCG) immunotherapy is the standard of care in treating non-muscle-invasive bladder cancer, yet its mechanism of action remains elusive. Both innate and adaptive immune responses have been implicated in BCG activity. Although prior research has indirectly demonstrated the importance of T cells and shown a rise in CD4 T cells in bladder tissue after BCG, T-cell subpopulations have not been fully characterized. We investigated the relationship between effector and regulatory T cells in an immune competent, clinically relevant rodent model of bladder cancer. Our data demonstrate that cancer progression in the -methyl--nitrosourea (MNU) rat model of bladder cancer was characterized by a decline in the CD8/FoxP3 ratio, consistent with decreased adaptive immunity. In contrast, treatment with intravesical BCG led to a large, transient rise in the CD4 T-cell population in the urothelium and was both more effective and immunogenic compared with intravesical chemotherapy. Whole-transcriptome expression profiling of posttreatment intravesical CD4 and CD8 T cells revealed minimal differences in gene expression after BCG treatment. Together, our results suggest that although BCG induces T-cell recruitment to the bladder, the T-cell phenotype does not markedly change, implying that combining T-cell-activating agents with BCG might improve clinical activity. .
Topics: Adaptive Immunity; Animals; BCG Vaccine; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Disease Models, Animal; Gene Expression Regulation, Neoplastic; Humans; Immunotherapy; Methylnitrosourea; Rats; Transcriptome; Urinary Bladder Neoplasms
PubMed: 28588015
DOI: 10.1158/2326-6066.CIR-16-0267