Spontaneous destructive periodontitis and skeletal bone damage in transgenic mice carrying a human shared epitope-coding HLA-DRB1 allele.

OBJECTIVE: Shared epitope (SE)-coding DRB1 alleles are associated with bone erosion in several diseases, including rheumatoid arthritis (RA) and periodontal disease (PD), but the underlying mechanism is unknown. We have recently identified the SE as an osteoclast-activating ligand. To better understand the biological effects of the SE in vivo, here we sought to determine whether it can facilitate spontaneous bone damage in naïve mice. METHODS: 3-month old naïve transgenic mice that carry the human SE-coding allele DRB1*04:01, or a SE-negative allele DRB1*04:02 were studied. Bone tissues were analysed by micro-CT, and the tooth-supporting tissues were studied by histology, immunohistochemistry and immunofluorescence. Serum biomarkers were determined by ELISA. RESULTS: Transgenic mice expressing the SE-coding DRB1*04:01 allele, but not mice carrying the SE-negative allele DRB1*04:02, showed spontaneous PD associated with interleukin (IL)-17 overabundance and periostin disruption. Mandibular bone volumetric and mineralisation parameters were significantly lower in SE-positive mice, and alveolar bone resorption was significantly increased in these mice. SE-positive mice also had more slender tibiae, and their marrow, cortical and total areas were lower than those of SE-negative mice. Additionally, significantly increased serum IL-17, tumour necrosis factor-α and osteoprotegrin levels were found in SE-positive mice, while their receptor activator of nuclear factor κ-B ligand levels were significantly lower. CONCLUSIONS: A human SE-coding allele increases the propensity to spontaneous bone-destructive periodontal inflammation and skeletal bone damage in transgenic mice. These findings provide new insights into the previously documented but poorly understood association of the SE with accelerated bone erosion in RA and several other human diseases.

Periodontal bacterial colonization in synovial tissues exacerbates collagen-induced arthritis in B10.RIII mice.

BACKGROUND: It has been previously hypothesized that oral microbes may be an etiological link between rheumatoid arthritis (RA) and periodontal disease. However, the mechanistic basis of this association is incompletely understood. Here, we investigated the role of periodontal bacteria in induction of joint inflammation in collagen-induced arthritis (CIA) in B10.RIII mice. METHODS: CIA-prone B10.RIII mice were infected orally with a polybacterial mixture of Porphyromonas gingivalis, Treponema denticola, and Tannerella forsythia for 24 weeks before induction of CIA. The ability of polybacterial mixture to colonize the periodontium and induce systemic response, horizontal alveolar bone resorption in infected B10.RIII mice was investigated. Arthritis incidence, severity of joint inflammation, pannus formation, skeletal damage, hematogenous dissemination of the infection, matrix metalloproteinase 3 (MMP3) levels, and interleukin-17 expression levels were evaluated. RESULTS: B10.RIII mice had gingival colonization with all three bacteria, higher levels of anti-bacterial immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies, significant alveolar bone resorption, and hematogenous dissemination of P. gingivalis to synovial joints. Infected B10.RIII mice had more severe arthritis, and higher serum matrix metalloproteinase 3 levels and activity. Histopathological analysis showed increased inflammatory cell infiltration, destruction of articular cartilage, erosions, and pannus formation. Additionally, involved joints showed had expression levels of interleukin-17. CONCLUSION: These findings demonstrate that physical presence of periodontal bacteria in synovial joints of B10.RIII mice with collagen-induced arthritis is associated with arthritis exacerbation, and support the hypothesis that oral bacteria, specifically P. gingivalis, play a significant role in augmenting autoimmune arthritis due to their intravascular dissemination to the joints.

Detection of ALDH1 activity in rabbit hepatic VX2 tumors and isolation of ALDH1 positive cancer stem cells.

BACKGROUND: Aldehyde dehydrogenase 1 (ALDH1) activity has been implicated in the therapeutic drug resistance of many malignancies and has been widely used as a marker to identify stem-like cells, including in primary liver cancer. Cancer stem cells (CSCs) are thought to play a crucial role in cancer progression and metastasis. In order to clarify the validity of the rabbit VX2 liver cancer model, we questioned if it expresses ALDH1 as a potential marker of CSCs. Hepatocellular carcinoma is a common malignancy worldwide and has poor prognosis. Most of the animal models used to study hepatocellular carcinoma are rodent models which lack clinical relevance. The rabbit VX2 model is a large animal model useful for preclinical and for developing drugs targeting cancer stem cells. MATERIALS AND METHODS: We used flow cytometry to identify rabbit VX2 liver tumor cells that express ALDH1A1 activity at a high level and confirmed the results with RT-PCR, immunohistochemical and western blot analyses. Further, mRNA and protein expression analysis of tumor samples also express the markers for stemness like klf4, oct3/4, CD44 and nanog as well as the differentiation marker α-fetoprotein. RESULTS: We used Aldefluor flow cytometry-based assay to identify cells with high ALDH1 activity in the rabbit VX2 liver cancer model. We used the brightest 4.39 % of the total cancer cell population in our study. We performed semi-quantitative as well as real time PCR to characterize the stemness derived from VX2 tumors and tissues from normal rabbit liver. We demonstrated that VX2 tumors have higher expression of cancer stem cell markers such as AlDH1A1 and CD44 in comparison to normal rabbit liver cells. Additionally, real time PCR analysis of the same samples using syber-green demonstrated the significant change (p > 0.05) in the expression of genes. We validated the gene expression of the stemness markers by performing western blot and immunofluorescence. We showed that cancer stem cell markers (AlDH1A1, CD44) and the differentiation marker α-fetoprotein were upregulated in VX2 tumor cells. The same extent of upregulation was observed in stemness markers (klf4, oct3/4 and nanog) in VX2 tumors in comparison to normal rabbit liver. CONCLUSION: The overall results of this study indicate that ALDH1 is a valid CSC marker for VX2 cancer. This finding suggests that the rabbit VX2 liver cancer model is useful in studying drug resistance in hepatocellular carcinoma and may be useful for basic and preclinical studies of other types of human cancer.

Inflammation and cancer: how friendly is the relationship for cancer patients?

Evidence has emerged in the last two decades that at the molecular level most chronic diseases, including cancer, are caused by a dysregulated inflammatory response. The identification of transcription factors such as NF-kappaB, AP-1 and STAT3 and their gene products such as tumor necrosis factor, interleukin-1, interleukin-6, chemokines, cyclooxygenase-2, 5 lipooxygenase, matrix metalloproteases, and vascular endothelial growth factor, adhesion molecules and others have provided the molecular basis for the role of inflammation in cancer. These inflammatory pathways are activated by tobacco, stress, dietary agents, obesity, alcohol, infectious agents, irradiation, and environmental stimuli, which together account for as much as 95% of all cancers. These pathways have been implicated in transformation, survival, proliferation, invasion, angiogenesis, metastasis, chemoresistance, and radioresistance of cancer, so much so that survival and proliferation of most types of cancer stem cells themselves appear to be dependent on the activation of these inflammatory pathways. Most of this evidence, however, is from preclinical studies. Whether these pathways have any role in prevention, progression, diagnosis, prognosis, recurrence or treatment of cancer in patients, is the topic of discussion of this review. We present evidence that inhibitors of inflammatory biomarkers may have a role in both prevention and treatment of cancer.

Models for prevention and treatment of cancer: problems vs promises.

Current estimates from the American Cancer Society and from the International Union Against Cancer indicate that 12 million cases of cancer were diagnosed last year, with 7 million deaths worldwide; these numbers are expected to double by 2030 (27 million cases with 17 million deaths). Despite tremendous technological developments in all areas, and President Richard Nixon's initiative in the 1974 "War against Cancer", the US cancer incidence is the highest in the world and the cancer death rate has not significantly changed in the last 50 years (193.9 per 100,000 in 1950 vs 193.4 per 100,000 in 2002). Extensive research during the same time, however, has revealed that cancer is a preventable disease that requires major changes in life style; with one third of all cancers assigned to Tobacco, one third to diet, and remaining one third to the environment. Approximately 20 billion dollars are spent annually to find a cure for cancer. We propose that our inability to find a cure to cancer lies in the models used. Whether cell culture or animal studies, no model has yet been found that can reproduce the pathogenesis of the disease in the laboratory. Mono-targeted therapies, till know in most cases, have done a little to make a difference in cancer treatment. Similarly, molecular signatures/predictors of the diagnosis of the disease and response are also lacking. This review discusses the pros and cons of current cancer models based on cancer genetics, cell culture, animal models, cancer biomarkers/signature, cancer stem cells, cancer cell signaling, targeted therapies, therapeutic targets, clinical trials, cancer prevention, personalized medicine, and off-label uses to find a cure for cancer and demonstrates an urgent need for "out of the box" approaches.

Radioprotective effects of Aloe vera leaf extract on Swiss albino mice against whole-body gamma irradiation.

The skin, being a cell-renewal system, is one of the first organs to be affected in total-body irradiation during radiotherapy. An attempt has been made in the present study to explore radiation-induced biochemical alterations caused by whole-body gamma irradiation and their modulation in Swiss albino mice by Aloe vera leaf extract (AVE). Mice were selected for this study from an inbreed colony and divided into four different groups: I (double-distilled water-treated group): considered as normal; II (Aloe vera-treated group): the animals were administered 1 g/kg body-wt/day Aloe vera leaf extract; III (radiation-treated group): the animals were exposed to 6 Gy gamma radiation at the dose rate of 0.96 Gy/min; and IV (combination group): animals were administered Aloe vera leaf extract continuously for 15 consecutive days, and on the 15th day they were irradiated to 6 Gy gamma radiation after 30 minutes of extract administration. The animals from the above groups were autopsied after 6 hours, 24 hours, and at 3, 7, 14, and 21 days of radiation. Biochemical estimations of DNA, lipid peroxidation, glutathione, catalase, and superoxide-dismutase were made. Total DNA, catalase, superoxide dismutase (SOD) activity in the skin, and glutathione (GSH) in the liver and blood significantly decreased compared to normal, but lipid peroxidation (LPO) in the liver and blood increased in the irradiated control group. In contrast, in experimental animals, DNA, catalase, and SOD in the skin and GSH in the liver and blood increased significantly, whereas LPO in the liver and blood decreased in comparison to irradiated control animals. Thus, Aloe vera leaf extract is found to have damage-resistant properties against radiation-induced biochemical alterations in Swiss albino mice.