Forward and reverse mutations in stages of cancer development.

BACKGROUND: Massive occurrences of interstitial loss of heterozygosity (LOH) likely resulting from gene conversions were found by us in different cancers as a type of single-nucleotide variations (SNVs), comparable in abundance to the commonly investigated gain of heterozygosity (GOH) type of SNVs, raising the question of the relationships between these two opposing types of cancer mutations. METHODS: In the present study, SNVs in 12 tetra sample and 17 trio sample sets from four cancer types along with copy number variations (CNVs) were analyzed by AluScan sequencing, comparing tumor with white blood cells as well as tissues vicinal to the tumor. Four published "nontumor"-tumor metastasis trios and 246 pan-cancer pairs analyzed by whole-genome sequencing (WGS) and 67 trios by whole-exome sequencing (WES) were also examined. RESULTS: Widespread GOHs enriched with CG-to-TG changes and associated with nearby CNVs and LOHs enriched with TG-to-CG changes were observed. Occurrences of GOH were 1.9-fold higher than LOH in "nontumor" tissues more than 2 cm away from the tumors, and a majority of these GOHs and LOHs were reversed in "paratumor" tissues within 2 cm of the tumors, forming forward-reverse mutation cycles where the revertant LOHs displayed strong lineage effects that pointed to a sequential instead of parallel development from "nontumor" to "paratumor" and onto tumor cells, which was also supported by the relative frequencies of 26 distinct classes of CNVs between these three types of cell populations. CONCLUSIONS: These findings suggest that developing cancer cells undergo sequential changes that enable the "nontumor" cells to acquire a wide range of forward mutations including ones that are essential for oncogenicity, followed by revertant mutations in the "paratumor" cells to avoid growth retardation by excessive mutation load. Such utilization of forward-reverse mutation cycles as an adaptive mechanism was also observed in cultured HeLa cells upon successive replatings. An understanding of forward-reverse mutation cycles in cancer development could provide a genomic basis for improved early diagnosis, staging, and treatment of cancers.

Integrated Network Pharmacology and in vivo Experimental Validation Approach to Explore the Potential Antioxidant Effects of Annao Pingchong Decoction in Intracerebral Hemorrhage Rats.

Background: Annao Pingchong decoction (ANPCD) is a traditional Chinese decoction which has definite effects on treating intracerebral hemorrhage (ICH) validated through clinical and experimental studies. However, the impact of ANPCD on oxidative stress (OS) after ICH remains unclear and is worth further investigating. Aim: To investigate whether the therapeutic effects of ANPCD on ICH are related to alleviating OS damage and seek potential targets for its antioxidant effects. Materials and Methods: The therapeutic candidate genes of ANPCD on ICH were identified through a comparison of the target genes of ANPCD, target genes of ICH and differentially expressed genes (DEGs). Protein-protein interaction (PPI) network analysis and functional enrichment analysis were combined with targets-related literature to select suitable antioxidant targets. The affinity between ANPCD and the selected target was verified using macromolecular docking. Subsequently, the effects of ANPCD on OS and the selected target were further investigated through in vivo experiments. Results: Forty-eight candidate genes were screened, in which silent information regulator sirtuin 1 (SIRT1) is one of the core genes that has antioxidant effects and ICH significantly affected its expression. The good affinity between 6 compounds of ANPCD and SIRT1 was also demonstrated by macromolecular docking. The results of in vivo experiments demonstrated that ANPCD significantly decreased modified neurological severity scoring (mNSS) scores and serum MDA and 8-OHdG content in ICH rats, while significantly increasing serum SOD and CAT activity, complicated with the up-regulation of ANPCD on SIRT1, FOXO1, PGC-1α and Nrf2. Furthermore, ANPCD significantly decreased the apoptosis rate and the expression of apoptosis-related proteins (P53, cytochrome c and caspase-3). Conclusion: ANPCD alleviates OS damage and apoptosis after ICH in rats. As a potential therapeutic target, SIRT1 can be effectively regulated by ANPCD, as are its downstream proteins.

Annao Pingchong decoction alleviate the neurological impairment by attenuating neuroinflammation and apoptosis in intracerebral hemorrhage rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Intracerebral hemorrhage (ICH) is a central nervous system disease that causes severe disability or death. Even though Annao Pingchong decoction (ANPCD), a traditional Chinese decoction, has been used clinically to treat ICH in China, its molecular mechanism remains unclear. AIM OF THE STUDY: To study whether the neuroprotective effect of ANPCD on ICH rats is achieved by alleviating neuroinflammation. This paper mainly explored whether inflammation-related signaling pathways (HMGB1/TLR4/NF-κB P65) plays a role in ANPCD treatment of ICH rats. MATERIALS AND METHODS: Liquid chromatography-tandem mass spectrometry was used to analyze the chemical composition of ANPCD. ICH models were established by injecting autologous whole blood into the left caudate nucleus of Sprague-Dawley (SD) rats. Modified neurological severity scoring (mNSS) was used to assess the neurological deficits. The levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and IL-6 were analyzed using enzyme-linked immunosorbent assay (ELISA). Pathological changes in the rat brains were observed using hematoxylin-eosin, Nissl, and TUNEL staining. The protein levels of HMGB1, TLR4, NF-κB p65, B-cell lymphoma 2 (Bcl-2), and Bcl-2-associated X protein (Bax) were measured by western blotting and immunofluorescence analysis. RESULTS: Ninety-three ANPCD compounds were identified, including 48 active plasma components. Treatment with ANPCD effectively improved the outcome, as observed by the neurological function scores analysis and brain histopathology. Our results showed that ANPCD exerts its anti-inflammatory effects by significantly downregulating the expression of HMGB1, TLR4, NF-κB p65, TNF-α, IL-1ß, and IL-6. ANPCD also exerted anti-apoptotic effects by significantly decreasing the apoptosis rate and Bax/Bcl-2 ratio. CONCLUSION: We found that ANPCD had neuroprotective effect in clinical work. Here, we also found that the action mechanism of ANPCD might be related to attenuate neuroinflammation and apoptosis. These effects were achieved by inhibiting the expression of HMGB1, TLR4 and NF-κB p65.