Hamster Buccal Pouch Carcinomas Research Articles

PLGA-Loaded Nedaplatin (PLGA-NDP) Inhibits 7,12-Dimethylbenz[a]anthracene (DMBA) Induced Oral Carcinogenesis via Modulating Notch Signaling Pathway and Induces Apoptosis in Experimental Hamster Model.

The present study is designed to evaluate the nanotherapeutic efficacy of prepared PLGA-loaded Nedaplatin (PLGA-NDP) against 7,12-dimethyl benz(a)anthracene (DMBA)-induced experimental oral carcinogenesis in hamster buccal pouch (HBP) model. The buccal pouch of golden Syrian hamsters was painted with 0.5% DMBA in liquid paraffin three times a week for 14 weeks, ultimately leading to the development of oral squamous cell carcinoma (OSCC). Oral administration of PLGA-NDP (preinitiation) and Cisplatin delivery (5 mg/kg b.wt) started 1 week before the carcinogen exposure and continued on alternative days. Post-administration of PLGA-NDP (5 mg/kg b.wt) started 2 days after carcinogen (DMBA) induction until the end of the experiment. After the 14th week, we observed that DMBA-painted hamsters exhibited tumor formation, morphological alterations, and well-differentiated OSSC in addition to the responsive molecular proteins during oral carcinogenesis. Furthermore, immunoblotting analysis demonstrated that PLGA-NDP inhibits Notch signaling, as evidenced by downregulation of Bcl-Xl, Bcl-2, p21, PGE2, HGF, and CXCL12 proteins, and upregulation of p53 and Bax. This apoptotic response is crucial for PLGA-NDP to induce apoptosis. In addition, RT-PCR results showed that PLGA-NDP nanoparticles play a downregulatory role in the therapeutic action of the notch signaling gene (Notch1, Notch 2, Hes1, Hey1, and Jagged1) at the mRNA transcription level in HBP carcinoma. Taken together, these data indicate that PLGA-NDP is a potent inhibitor of oral carcinogenesis and the expansion of cells that specifically target the Notch signaling pathway indicates that obstructing Notch signaling could potentially serve as a new and innovative therapeutic approach for oral squamous cell carcinoma (OSCC).

Therapeutic efficacy of axitinib as anti-angiogenesis and nivolumab as immunotherapy on DMBA induced hamster buccal pouch carcinoma utilizing vascular endothelial growth factor

Therapeutic efficacy of axitinib as anti-angiogenesis and nivolumab as immunotherapy on DMBA induced hamster buccal pouch carcinoma utilizing vascular endothelial growth factor

Tumour volume and s-phase fraction analysis in induced hamster buccal pouch carcinoma utilizing cetuximab immunotherapy.

Tumour volume and s-phase fraction analysis in induced hamster buccal pouch carcinoma utilizing cetuximab immunotherapy.

The Neem Limonoid Nimbolide Modulates Key Components of the DNA Damage Response Signalling in Cellular and Animal Models of Oral Squamous Cell Carcinoma

Deregulated DNA damage response (DDR) network is implicated in cancer progression and therapy resistance. The present study was designed to investigate whether nimbolide, an anticancer neem limonoid, targets key components of the DDR signalling pathway in cellular and animal models of oral squamous cell carcinoma (OSCC). OSCC cells (SCC-4 and SCC-9), 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinoma model, chemoresistant OSCC patient-derived xenograft (PDX) model established in athymic nude mice, and tissue sections from patients with oral premalignant/malignant disease were used for the study. Key molecules that orchestrate the DDR, including the MRN complex, ATM, DNA-PKcs, H2AX, and p53, were analysed by qRT-PCR, immunoblotting, immunofluorescence, and immunohistochemistry. Cell proliferation and apoptosis indices were evaluated. Nimbolide significantly reduced 8-oxodG levels, expression of MRN, ATMS1891, and γ-H2AX, with an increase in p-p53S15 in OSCC cells as well as in the HBP model. Nimbolide potentiated the effect of KU-55933 in ATM inhibition. In the PDX model, nimbolide suppressed tumor formation, stimulated DDR and apoptosis, inhibited cell proliferation, and enhanced sensitivity to cisplatin. Analysis of p-ATM expression revealed a significant increase during the sequential progression of hamster and human OSCC. This study provides compelling evidence that nimbolide functions as a DDR inhibitor in cellular and hamster OSCC models and as a DDR activator in the PDX model primarily by targeting ATM. Small molecules like nimbolide that modulate DDR are of immense benefit in cancer therapy. The study has also unveiled p-ATM as a promising biomarker of tumour progression in human OSCCs.

Assessment of DNA ploidy on experimentally induced hamster buccal pouch carcinoma (Histological and flow cytometric studies)

Assessment of DNA ploidy on experimentally induced hamster buccal pouch carcinoma (Histological and flow cytometric studies)

Therapeutic potential of epigallocatechin-3-gallate on cancer stem cells of DMBA induced hamster buccal pouch carcinoma

Therapeutic potential of epigallocatechin-3-gallate on cancer stem cells of DMBA induced hamster buccal pouch carcinoma

Therapeutic efficacy of time-dependent cetuximab on experimentally induced hamster buccal pouch carcinoma

Purpose: This study aimed to determine the cetuximab therapeutic potential in the time-dependent fashion in experimentally induced hamster buccal pouch (HBP) carcinoma. Material and methods: Forty Syrian male hamsters were classified into four equal groups (G) of ten each. GI: The animals remained untreated to act as negative controls. The right pouches of animals in GII, GIII, and GIV were painted three times a week for 14 weeks (w)s with 7,12-dimethylbenz (a) anthracene (DMBA). GII: No additional treatment was administered. While, the animals in GIII and GIV were treated differently. Those in GIII received cetuximab intraperitoneally (IP) three intervals a week for three (w)s, whereas those in GIV received cetuximab IP three intervals for six (w)s. After the end of the experiment, the gross observations were made, and blood samples (2ml) were withdrawn from the inner canthus of the eye for analysis of whole white blood cells (WBCs) and oxidative stress markers [glutathione (GSH) and malondialdehyde (MDA) levels]. All pouches were surgically bisected for preparation for Hematoxylin and Eosin (H&E) stain and immunohistochemistry (IHC) stain using epidermal growth factor receptor (EGFR). Other fresh tissue was used for DNA detection through a flow cytometry (FCM) test.

Therapeutic efficacy of intraperitoneally injected curcumin on experimentally induced hamster buccal pouch carcinoma

Abstract: The aim of the present study was directed to investigate the therapeutic efficacy of intraperitoneally injected curcumin as anticarcinogenic treatment modality on 7,12-dimethylbenz[a]anthracene (DMBA) induced hamster buccal pouch(s) (HBP(s)) carcinoma. Subjects and methods: Thirty Syrian male hamsters, five weeks old, weighing 90-130g were divided into three group(s) (G(s)): A, B, and C. G-A (negative control): HBP mucosa of 10 animals were painted with liquid paraffin alone three times a week for 14 weeks. G-B (positive control): 10 animals were painted with 0.5% DMBA in paraffin oil 3 times a week for 14 weeks. Then animals were left untreated for 6 weeks after DMBA painting. G-C (curcumin treated group): 10 animals were injected intraperitoneally with curcumin 80 mg/kg body weight every day for 6 weeks following DMBA-treatment. The assessment was based on the gross observation, histological and immunohistochemical examination utilizing anti-apoptotic Bcl-2 antibody. Results: Gross observation revealed variation in reduction of the tumor size in the treated group (G-C) compared to that observed in group G-B. Histopathological findings revealed variations between G-B and G-C groups. Immunohistochemical results revealed that Bcl-2 expression has a variability in the area % throughout the groups used. Groups G-A, G-B, and G-C were 7.78%, 75.51%, and 41.65% respectively. Conclusion: The therapeutic effect of intraperitoneally injected curcumin resulted in promising outcome in regression of DMBA induced HBP carcinoma.

Function of gold nanoparticles in oral cancer beyond drug delivery: Implications in cell apoptosis

Gold nanoparticles (AuNPs) are used to deliver drugs and therapeutic small molecule inhibitors to cancer cells. Evidence shows that AuNPs coated with nuclear localization sequence can cross the nuclear membrane and induce cellular apoptosis. To determine the therapeutic role of AuNPs, we compared two nanoconstructs conjugated to doxorubicin (DOX) through pH-sensitive and pH-resistant linkers. We tested DOX nanoconjugates' cytotoxicity, cellular and nuclear uptake in oral squamous cell carcinoma cell line. Furthermore, we evaluated the therapeutic effect of pH-sensitive and pH-resistant DOX bioconjugates in hamster buccal pouch carcinoma model. Our data indicate that pH-resistant and pH-sensitive DOX-nanoconjugates were equally localized in cancer cells, but the pH-resistant DOX nanoparticles were more localized in the nuclei inducing a 2-fold increase in the apoptotic effect compared with the pH-sensitive DOX nanoparticles. Our in vivo results show significantly higher tumor shrinkage and survival rates in animals treated with DOX pH-resistant AuNPs compared with pH-sensitive ones. Our findings suggest that AuNPs enhance the cytotoxic effect against cancer cells in addition to acting as drug carriers. DOX pH-resistant AuNPs enhanced accumulation of AuNPs in cancer cells' nuclei inducing a significant cellular apoptosis which was confirmed using in vitro and in vivo experiments without deleterious effects on blood cell count.

CHEMOPREVENTIVE EFFECT OF GREEN TEA AND CURCUMIN IN INDUCED ORAL SQUAMOUS CELL CARCINOMA: AN EXPERIMENTAL STUDY

Introduction: Oral squamous cell carcinoma (OSCC) has the highest mortality rates among all carcinomas and is the most common head and neck cancer. Several natural compounds and micronutrients have been under investigation for their efficacy in head and neck cancer chemoprevention. Green tea contains various cancer preventive catechins that have a role in preventing cancer recurrence in various organs in humans. Curcumin, a natural polyphenol, is one of the most investigated biomolecules from Mother Nature. Curcumin has been shown to exert significant growth inhibitory effect on oral precancerous and carcinoma cell lines, and the effect is synergistic with epigallocatechin gallate, the most abundant polyphenol in tea. Objectives: To assess the chemopreventive effects of green tea and curcumin through induction of hamster buccal pouch carcinoma by using an apoptotic marker and compare their effect each alone and in combination. Materials and methods: Squamous cell carcinoma was chemically induced in fifty Syrian golden hamsters divided into 5 groups (10 each). The first group was used as normal control group. The second group received the carcinogenic agent only. The other three groups received green tea, curcumin and combination of both, respectively. Results:Normal control group (A) revealed neither pathological nor inflammatory changes in the buccal pouch with 1.72%of the cells underwent apoptosiswhile the cells of positive control group (B) resulted in 11.57% apoptosis. In the study groups, treatment of the cells with green tea (C), curcumin (D) and both of them (E) resulted in 82.22 %, 78.91%, 96.63% apoptosis respectively. The fluorescent image by confocal laser scanning in group B showed increase of the red fluorescence in the nucleus and cytoplasm of the squamous cell carcinoma cells indicating high proliferating cells while the fluorescent image of group C, D and E showed decrease of the red florescence in the nuclei of the squamous cell carcinoma cells indicating low proliferation. Conclusions: Green tea and curcumin have a significant chemopreventive effect against oral carcinogenesis and the combination of both agents has a better effect.

Stem cell exosomes inhibit angiogenesis and tumor growth of oral squamous cell carcinoma

Recently, exosomes secreted by menstrual mesenchymal stem cells have been identified as inhibitory agents of tumor angiogenesis and modulators of the tumor cell secretome in prostate and breast cancer. However, their direct effect on endothelial cells and paracrine mediators have not yet been investigated. Using a carrier-based cell culture system to test the scalability for exosome production, we showed that different types of endothelial cells present specific kinetics for exosomes internalization. Exosome-treatment of endothelial cells increased cytotoxicity and reduced VEGF secretion and angiogenesis in a dose-dependent manner. Using the hamster buccal pouch carcinoma as a preclinical model for human oral squamous cell carcinoma, we demonstrated a significant antitumor effect of intra-tumoral injection of exosomes associated with a loss of tumor vasculature. These results address up-scaling of exosome production, a relevant issue for their clinical application, and also assess menstrual stem cell exosomes as potential anti-angiogenic agents for the treatment of neoplastic conditions.

Chemoprevention efficacy of indole-3-Carbinol in experimentally induced hamster buccal pouch carcinogenesis

Abstract: The aim of the present study was directed to investigate the effect of indole-3-carbinol (I3C) as a new chemopreventive modality in experimentally induced hamster buccal pouch (HBP) carcinogenesis. Material and methods: Twenty five golden Syrian male hamsters five weeks old, weighting 80-120 gs were used as experimental animals and divided into three group(s) (G(S)) (G1, G2 and G3): G1 (negative control): 5 animals were left untreated. G2: (dimethylbenz[a]anthracene (DMBA) painting-HBP group): 10 animals, their right buccal pouches were painted with 0.5% DMBA in paraffin oil 3 times a week and divided into two subgroups, G2A and G2B (G2A which included 5 animals and painted for 8 weeks, G2B which included 5 animals and painted for 14 weeks. G3 (I3C chemoprevention group): 10 hamsters were included in this group were received I3C given by the oral route using a specific vehicle one week before, as well as during the application of DMPA on alternative days for 8 weeks (G3A) and for 14 weeks (G3B). After termination of the experiment, all animals were sacrificed, and the buccal mucosa was excised and fixed in 10% neutral buffered formalin, routinely processed and embedded in paraffin blocks for preparation in order to be examined histologically and immunohistochemically and then statistical analysis based on these examinations was done Results: Gross observations revealed variable features in (G2 and G3) compared to that observed in group G1ranging from normal and smooth surface to fungating tumor masses of large sizes. Histopathological findings revealed variations among chemoprevention groups ranging from normal epithelial layers to epithelial dysplasia to squamous cell carcinoma with invading nests and pearls. Immunohistochemical (IHC) results, regarding Bax expression, revealed variability in the area percentage throughout the groups used. At 8 weeks, area percentages of G1, G2A, and G3A were (39.83%, 22.00 %, and 30.47 %) respectively, and at 14 weeks were (39.83%, 10.45 % and 26.19%) respectively. Bcl2 expressions also had variability in the area percentage throughout the same groups at 8 weeks were (7.05 %, 43.09 % and 18.15 %) respectively, while at 14 weeks they were (7.05 %, 75.21 %and 43.76 %) respectively. Conclusion: I3C is considered as a promising chemotherapeutic agent in prevention of induced HBP carcinogenesis (epithelial dysplasia & invasive carcinoma) and proved beneficial role in improving the outcome by modulating apoptosis and proliferation throughout the process of carcinogenesis.Keywords: HBP carcinoma, indole-3-carbinol, Bax, Bcl2.

Gramine inhibits angiogenesis and induces apoptosis via modulation of TGF-β signalling in 7,12 dimethylbenz[a]anthracene (DMBA) induced hamster buccal pouch carcinoma

BackgroundTransforming growth factor-β (TGF-β) and its receptors are considered as a novel target in cancer chemotherapy. Gramine, an indole alkaloid, possesses various pharmacological properties including antiproliferative and anticancer. However, the anti-angiogenic property remains unexplored. PurposeThe present study was designed to evaluate the anti-angiogenic and apoptosis induction properties of gramine through inhibiting TGF-β on DMBA induced oral squamous cell carcinoma (OSCC) in the hamster buccal pouch (HBP). MethodsThe effects of gramine on TGF-β signalling in DMBA induced carcinogenic events such as angiogenesis and apoptosis were analysed by studying the mRNA expression using RT-PCR, protein expression by western blot and histopathological analysis using haematoxylin and eosin (H & E) staining. ResultsGramine significantly inhibited phosphorylation and nuclear translocation of Smad2 and Smad4 by blocking activity of the TGFβ-RII, RI and activation of inhibitory Smad7. Gramine inhibited angiogenic markers such as MMP-2, MMP-9, HIF-1α, VEGF, and VEGF-R2 as well as increased TIMP-2 expression. Furthermore, gramine induced apoptosis in DMBA induced tumour bearing animals by up regulating the pro apoptotic proteins Bax, cytochrome C, apaf-1, caspase-9 caspase-3 and PARP. ConclusionIn this study, we clearly demonstrated that gramine treatment diminishes angiogenesis and induces apoptosis in hamster buccal pouch (HBP) carcinogenesis by modulating TGF-β signals.

Synergistic effect between cetuximab and curcumin on experimentally induced hamster buccal pouch carcinoma

Synergistic effect between cetuximab and curcumin on experimentally induced hamster buccal pouch carcinoma

Evaluation of caspase 9 expression in induced hamster buccal pouch carcinoma

Evaluation of caspase 9 expression in induced hamster buccal pouch carcinoma

Kras gene mutation analysis in experimentally induced hamster buccal pouch carcinoma

Kras gene mutation analysis in experimentally induced hamster buccal pouch carcinoma

Diosmin induce apoptosis through modulation of STAT-3 signaling in 7,12 dimethylbenz(a)anthracene induced harmster buccal pouch carcinogenesis

Diosmin is naturally found flavanoid in many citrus fruits known to have anti-inflammatory, antihyperglycemic, antioxidant and antimutagenic properties. Effects of Diosmin on IL-6/STAT-3 expression in hamster buccal pouch carcinogenesis remain unclear. Alterations in many genes encode crucial proteins, which regulate cell proliferation, differentiation and apoptosis have been implicated in oral cancer. In the present study, we investigated the effect of dietary Diosmin on IL-6/STAT-3 signaling in the 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis by examining the protein expression of IL-6/STAT-3 and its related genes. Immunoblotting and immunohistochemical analyses revealed that Diosmin (100mg/kgb.w) supplement inhibits key events in signaling especially STAT-3 phosphorylation and subsequent nuclear translocation. Results revealed that inhibition of proliferation and angiogenesis is associated with regulation of the STAT-3 pathway; where Diosmin prevents phosphorylation of JAK-1 which was ascend by IL-6, thereby inhibiting STAT-3 phosphorylation. Consequently, an imbalance in the Bax/Bcl-2 ratio triggered the caspase cascade in favor of apoptosis. Transmission electron microscopic studies proved the effect of Diosmin on ultrastructural changes. Finally our results provide significant evidence that Diosmin prevents the development and progression of HBP carcinomas through the inhibition of IL-6/STAT-3 signaling and its downstream events. Thus, Diosmin functions as a potent inhibitor of tumor development and progression by targeting IL-6/STAT-3 signaling may be an ideal candidate for cancer chemoprevention.

Blueberry inhibits invasion and angiogenesis in 7,12-dimethylbenz[a]anthracene (DMBA)-induced oral squamous cell carcinogenesis in hamsters via suppression of TGF-β and NF-κB signaling pathways

Aberrant activation of oncogenic signaling pathways plays a pivotal role in tumor initiation and progression. The purpose of the present study was to investigate the chemopreventive and therapeutic efficacy of blueberry in the hamster buccal pouch (HBP) carcinogenesis model based on its ability to target TGF-β, PI3K/Akt, MAPK and NF-κB signaling and its impact on invasion and angiogenesis. Squamous cell carcinomas were induced in the HBP by 7,12-dimethylbenz[a]anthracene (DMBA). The effect of blueberry on the oncogenic signaling pathways and downstream events was analyzed by quantitative real-time PCR and immunoblotting. Experiments with the ECV304 cell line were performed to explore the mechanism by which blueberry regulates angiogenesis. Blueberry supplementation inhibited the development and progression of HBP carcinomas by abrogating TGF-β and PI3K/Akt pathways. Although blueberry failed to influence MAPK, it suppressed NF-κB activation by preventing nuclear translocation of NF-κB p65. Blueberry also modulated the expression of the oncomiR miR-21 and the tumor suppressor let-7. Collectively, these changes induced a shift to an anti-invasive and anti-angiogenic phenotype as evidenced by downregulating matrix metalloproteinases and vascular endothelial growth factor. Blueberry also inhibited angiogenesis in ECV304 cells by suppressing migration and tube formation. The results of the present study suggest that targeting oncogenic signaling pathways that influence acquisition of cancer hallmarks is an effective strategy for chemointervention. Identification of modulatory effects on phosphorylation, intracellular localization of oncogenic transcription factors and microRNAs unraveled by the present study as key mechanisms of action of blueberry is critical from a therapeutic perspective.

Nimbolide, a neem limonoid inhibits Phosphatidyl Inositol-3 Kinase to activate Glycogen Synthase Kinase-3β in a hamster model of oral oncogenesis.

Glycogen synthase kinase-3β (GSK-3β), a serine/threonine kinase is frequently inactivated by the oncogenic signalling kinases PI3K/Akt and MAPK/ERK in diverse malignancies. The present study was designed to investigate GSK-3β signalling circuits in the 7,12-dimethylbenz[a]anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis model and the therapeutic potential of the neem limonoid nimbolide. Inactivation of GSK-3β by phosphorylation at serine 9 and activation of PI3K/Akt, MAPK/ERK and β-catenin was associated with increased cell proliferation and apoptosis evasion during stepwise evolution of HBP carcinomas. Administration of nimbolide inhibited PI3K/Akt signalling with consequent activation of GSK-3β thereby inducing trafficking of β-catenin away from the nucleus and enhancing the expression of miR-126 and let-7. Molecular docking studies confirmed interaction of nimbolide with PI3K, Akt, ERK and GSK-3β. Furthermore, nimbolide attenuated cell proliferation and induced apoptosis as evidenced by increased p-cyclin D1Thr286 and pro-apoptotic proteins. The present study has unravelled aberrant phosphorylation as a key determinant for oncogenic signalling and acquisition of cancer hallmarks in the HBP model. The study has also provided mechanistic insights into the chemotherapeutic potential of nimbolide that may be a useful addition to the armamentarium of natural compounds targeting PI3K for oral cancer treatment.

The Hamster Buccal Pouch Model of Oral Carcinogenesis.

The hamster buccal pouch (HBP) carcinogenesis model is one of the most well-characterized animal tumor models used as a prelude to investigate multistage oral carcinogenesis and to assess the efficacy of chemointervention. Hamster buccal pouch carcinomas induced by 7,12-dimethylbenz[a]anthracene (DMBA) show extensive similarities to human oral squamous cell carcinomas. The HBP model offers a number of advantages including a simple and predictable tumor induction procedure, easy accessibility for examination and follow-up of lesions, and reproducibility. This model can be used to test both chemopreventive and chemotherapeutic agents.