| PurposeThe etiology and mechanism of tumorigenesis is currently unclear, the etiology-based tumor-bearing animal models for screening anti-tumor drugs are absent, and mechanism-dependent effective prevention and treatment protocols are also unavailable. Therefore, we suggest here, for the first time, a hypothesis of’inflammation-triggered nitric oxide (NO)-driven hypoxia-induced tumorigenesis’. Considering our previous results of the immune stimuli provoking NO burst in the investigation of articular synovitis, and the similarity of symptomatic manifestations and pathological characteristics of primary synovial pathogenesis with tumor-like angiogenesis and hyperplasia, we try to induce hypoxia in the local mouse tissues using the exogenous NO producer sodium nitroprusside (SNP) and the endogenous NO generators collagen Ⅱ(CⅡ), complete Freund’s adjuvant (CFA), and liposaccharide (LPS). The mouse models with NO-induced tumor-like hypodermal and synovial hyperplasia are separately established, the possibility of’hypoxia-induced tumorigenesis’is preliminarily verified, and the effect of selective anti-tumor candidate drugs that suppress tumor-like in situ hyperplasia is evaluated.Material and MethodsThis study is attempting to establish a mouse model with tumor-like in situ hypodermal hyperplasia by tying the dorsal fur to restrict the blood supply and injecting into it with SNP, CII, or CFA, in a singular component or in combinations; and also trying to establish a mouse model with synovial tumor-like in situ hyperplasia by injecting into the articular cavity with SNP, CFA, or LPS. Using those non-grafted tumor models, pathogenetic research will be carried out from the aspects of morphology, histochemistry, immune histochemistry, blood hypoxia indicators, pro-inflammatory cytokine secretion dynamics, protein nitrosylation extents, and hypoxia responsive gene expression profiles. Upon the mouse models with synovial tumor-like hyperplasia, the therapeutic effects of anti-tumor candidate drugs including two kinds of the phytochemical components, betulilic acid and artemisinin (the soluble derivative artesunate), and one kind of the Traditional Chinese Medicine Patent Prescription, Xuefu Zhuyu tablet, will be evaluated and compared.ResultsEither the SNP-released exogenous NO or the C Ⅱ, CFA, LPS-provoked endogenous NO enhances the localized angiogenesis and hyperplasia within mouse hypodermal or synovial tissues, hence tentatively demonstrating that NO is a crucial effector leading to tumor-like hyperplasia. During those processes, elevation of the blood NO level is accompanied with decline of the blood oxygen saturation percentage (SpO2) in the hypoderm or articulate, suggesting that NO is a direct reason for inducing local tissue hypoxia. At the same time, it is found that SNP and CⅡ-CFA induce the expression of hypoxia responsive genes, such as mitochondrial uncoupled protein2(UCP2) gene and several cytokine (IL-1α, IL-6, NOTCH1) genes. While SNP down-regulates NOS2(iNOS) gene, CⅡ-CFA up-regulates NOS2gene. It is also noted that both endogenous and exogenous NO significantly up-regulate angiogenesis-related hypoxia inducible factor1α (HIF-1α) and vascular endothelial growth factor (VEGF). Consequently, the present study has reasonably associated NO with such processes as hypoxia, angiogenesis, and hyperplasia, and also successfully established the mouse models with pathogenesis-based tumor-like hyperplasia.Upon using the mouse model with synovial tumor-like hyperplasia, we have evaluated the anti-hyperplasia effects and investigated the action mechanisms of several anti-tumor candidate drug monomers and compounds, including betulilic acid, artesunate, and Xuefu Zhuyu tablet. The results indicate that all tested drugs exhibit the remarked effects combating tumor-like hyperplasia, in which betulilic acid blocks hyperplasia mainly through suppressing the transcriptional factor HIF-1α-induced VEGF expression; artesunate blocks hyperplasia mainly through inhibiting NO production; and Xuefu Zhuyu table blocks hyperplasia principally through activating blood circulation to dissipate blood stasis and thereby mitigating local tissue hypoxia. The effects against tumor-like hyperplasia of those above drugs have been evaluated from tumor morphology, histopathology (histochemistry and immune histochemistry) and blood hypoxia indicators (NO and SpO2).ConclusionsThis study has preliminarily verified the hypothesis of NO-mediated hypoxia accelerating tumor-like hyperplasia, and connected tumorigenesis together with the critical steps such as immune activation, NO burst, hypoxia induction, angiogenesis, and hyperplasia. We have successfully established the non-grafted mouse models with in situ tumor-like hyperplasia, by which the effects and mechanisms against tumor-like hyperplasia of several anti-tumor candidate drugs have been quickly evaluated. |
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