Fucoidan as an Autophagy Regulator: Mechanisms and Therapeutic Potentials for Cancer and Other Diseases.

Fucoidan, a natural polysaccharide with a variety of classical bioactivities mainly sourced from brown algae, has been extensively studied owing to its favorable pharmacological effects, including anti-inflammatory, anti-tumor, anticoagulant and liver protection. Recently it has been found to play a regulatory role in the processes of autophagy. Autophagy is an important cellular process that effectively protects cells and organisms from stimulating factors such as nutrient deficiency, low cellular ATP levels, metabolic stress, growth factor deprivation and hypoxic conditions. In recent years, many studies have shown that fucoidan can treat human diseases by regulating autophagy process though cell signaling pathways. In this review, we summarize the latest progress in the discovery of natural autophagy regulator of fucoidan for the therapeutic application in cardiac diseases, cancers and liver diseases, aiming to provide the new pharmacological application that fucoidan may treat human diseases by regulating autophagy. Furthermore, we look forward to seeing more diseases that would be treated by autophagy modulator of fucoidan and the discovery of more elaborate autophagy regulation mechanism.

Inhibition of fucoidan on breast cancer cells and potential enhancement of their sensitivity to chemotherapy by regulating autophagy.

Fucoidan is a marine-origin sulfated polysaccharide that has gained attention for its anticancer activities. However, the inhibitory effect of fucoidan on breast cancers by regulating autophagy and its mechanism are not clear, and the chemotherapeutic sensitization of fucoidan is largely unknown. In the present study, the anticancer potential of fucoidan was revealed in MCF-7 and MDA-MB-231 cells. Additionally, we also studied the chemotherapeutic sensitization of fucoidan by combining chemotherapeutic drugs doxorubicin (ADM) and cisplatin (DDP) with fucoidan on breast cancer cells. In the two kinds of human breast cancer cells, cell viability was determined by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Apoptosis was examined with flow cytometry. Transfection assay was used to examine autophagy flow. Western blot was used to examine the expressions of related proteins. Results suggested that fucoidan could induce autophagy and might enhance the sensitivity of breast cancer cells to chemotherapeutic drugs. Mechanistically, fucoidan induced autophagy in breast cancer cells by down-regulating m-TOR/p70S6K/TFEB pathway. In conclusion, our research revealed that fucoidan could induce autophagy of breast cancer cells by mediating m-TOR/p70S6K/TFEB pathway, thus inhibiting tumor development. Furthermore, fucoidan might enhance the sensitivity of breast cancer cells to ADM and DDP, and this enhancement was related to autophagy.

Metabolic regulation mechanism of fucoidan via intestinal microecology in diseases.

The intestinal microecology is an extremely complex ecosystem consisting of gut microbiota, intestinal mucosa and the intestinal immune system. The intestinal microecology performs several important functions and is considered to be an essential 'organ' because it plays an important role in regulating human metabolism. Fucoidan contains a large amount of fucose and galactose residues, as well as various other neutral and acidic monosaccharides. Fucoidan particularly effects tumors, inflammatory bowel disease, diabetes and obesity by repairing intestinal mucosal damage and improving the intestinal microecological environment. It has been proposed that fucoidan could be used as a prebiotic agent for pharmaceutical and functional foods. In this review, we elucidate the potential mechanisms of the metabolic regulation of fucoidan with respect to the intestinal microecology of diseases. © 2021 Society of Chemical Industry.

The molecular mechanism and evolutionary divergence of caspase 3/7-regulated gasdermin E activation.

Caspase (CASP) is a family of proteases involved in cleavage and activation of gasdermin, the executor of pyroptosis. In humans, CASP3 and CASP7 recognize the same consensus motif DxxD, which is present in gasdermin E (GSDME). However, human GSDME is cleaved by CASP3 but not by CASP7. The underlying mechanism of this observation is unclear. In this study, we identified a pyroptotic pufferfish GSDME that was cleaved by both pufferfish CASP3/7 and human CASP3/7. Domain swapping between pufferfish and human CASP and GSDME showed that the GSDME C-terminus and the CASP7 p10 subunit determined the cleavability of GSDME by CASP7. p10 contains a key residue that governs CASP7 substrate discrimination. This key residue is highly conserved in vertebrate CASP3 and in most vertebrate (except mammalian) CASP7. In mammals, the key residue is conserved in non-primates (e.g., mouse) but not in primates. However, mouse CASP7 cleaved human GSDME but not mouse GSDME. These findings revealed the molecular mechanism of CASP7 substrate discrimination and the divergence of CASP3/7-mediated GSDME activation in vertebrate. These results also suggested that mutation-mediated functional alteration of CASP probably enabled the divergence and specialization of different CASP members in the regulation of complex cellular activities in mammals.

Cell death is essential for an organism to develop and survive as it plays key roles in processes such as embryo development and tissue regeneration. Cell death is also an important form of defence during an infection. A form of programmed cell death known as pyroptosis can be induced in infected cells, which helps to kill the infectious agent as well as alert the immune system to the infection. Pyroptosis is driven by Gasdermin E, a protein made up of two domains. At one end of the protein, the 'N-terminal' domain punctures holes in cell membranes, which can lead to cell death. At the other end, the 'C-terminal' domain inhibits the activity of the N-terminal domain. A family of proteins called caspases activate Gasdermin E by cleaving it, which releases the N-terminal domain from the inhibitory C-terminal domain. In humans, two caspases known as CASP3 and CASP7 recognize a specific sequence of amino acids ­ the building blocks of proteins ­ in Gasdermin E. However, only CASP3 is able to cleave the protein. After discovering that, unlike in humans, pufferfish Gasdermin E can be cleaved by both CASP3 and CASP7, Xu et al. wanted to investigate the underlying mechanisms behind this difference. Swapping the domains of human and pufferfish Gasdermin E and creating different versions of CASP7 revealed that the C-terminal domain of Gasdermin E and a single amino acid in CASP7 determine whether cleavage is possible. Interestingly, the key amino acid sequence required for cleavage by CASP7 is present in most vertebrate CASP3 and CASP7 proteins. However, it is absent in most mammalian CASP7. The findings of Xu et al. suggest that the different activity of human CASP7 and CASP3 is driven by a single amino acid mutation. This change likely played an important role in the process of different CASP proteins evolving to regulate different cellular activities in mammalian cells. This knowledge will be useful for future studies on the evolution and specialization of other closely related proteins.

Knockdown of DJ-1 Exacerbates Neuron Apoptosis Induced by TgCtwh3 through the NF-κB Pathway.

Mutations or loss of function of DJ-1 and Toxoplasma gondii (T. gondii) infection has been linked to neurodegenerative diseases, which are often caused by oxidative stress. However, the relationship between DJ-1 and T. gondii infection is not yet fully understood. Therefore, this study aimed to investigate the expression of DJ-1 in the hippocampus tissue of mice or in HT22 infected with T. gondii Chinese 1 genotype Wh3 strain (TgCtwh3) and the effect of DJ-1 knockdown on neuronal apoptosis induced by TgCtwh3 tachyzoite, as well as the underlying mechanism at the cellular and molecular level. Firstly, we detected DJ-1 protein expression and cell apoptosis in the hippocampal tissue of mice infected by TgCtwh3. Then, we examined DJ-1 expression and apoptosis in HT22 challenged with TgCtwh3. Finally, we evaluated the apoptosis in HT22 with DJ-1 knockdown which was infected with TgCtwh3 and assayed the expression of NF-κBp65 and p-NF-κBp65. Our results showed that DJ-1 expression was reduced and neurons underwent apoptosis in the hippocampus of mice infected with TgCtwh3 tachyzoites. Additionally, the knockdown of DJ-1 followed by infection with TgCtwh3 tachyzoites led to increased apoptosis in HT22 cells through the NF-κB signaling pathway. Therefore, this study suggests that DJ-1 is an important target for preventing apoptosis caused by T. gondii TgCtwh3.

Time-lapse live-cell imaging of pyroptosis by confocal microscopy.

Pyroptosis is a type of regulated necrosis executed by gasdermin. Osmotic cell swelling and membrane perforation are the key features of pyroptosis. This protocol presents time-lapse imaging of morphological changes during pyroptosis using a confocal microscope. We describe the step-by-step ectopic expression of gasdermin, cell staining with nuclear and membrane probes, and visualization of pyroptosis by time-lapse imaging. This protocol is applicable to monitoring pyroptosis in various situations. For complete details on the use and execution of this protocol, please refer to Qin et al. (2023).1.

Pyroptotic gasdermin exists in Mollusca and is vital to eliminating bacterial infection.

Gasdermin (GSDM) is a family of proteins that execute pyroptosis in vertebrate. In invertebrate, pyroptotic GSDM was documented only in coral. Recent studies identified abundant GSDM structural homologs in Mollusca, but their functions are unclear. Herein, we report a functional GSDM from Pacific abalone Haliotis discus (HdGSDME). HdGSDME is specifically activated by abalone caspase 3 (HdCASP3) cleavage at two distinct sites, generating two active isoforms with pyroptotic and cytotoxic activities. HdGSDME possesses evolutionarily conserved residues that proved to be essential to the N-terminal pore-formation and C-terminal auto-inhibition capacities. Bacterial challenge activates the HdCASP3-HdGSDME pathway and induces pyroptosis and extracellular traps in abalone. Blockage of the HdCASP3-HdGSDME axis promotes bacterial invasion and host mortality. Collectively, this study reveals the existence of functionally conserved and yet distinct-featured GSDM in Mollusca and provides insights into the function and evolution of invertebrate GSDM.

Erythropoietin inhibits ferroptosis and ameliorates neurological function after spinal cord injury.

Ferroptosis is one of the critical pathological events in spinal cord injury. Erythropoietin has been reported to improve the recovery of spinal cord injury. However, whether ferroptosis is involved in the neuroprotective effects of erythropoietin on spinal cord injury has not been examined. In this study, we established rat models of spinal cord injury by modified Allen's method and intraperitoneally administered 1000 and 5000 IU/kg erythropoietin once a week for 2 successive weeks. Both low and high doses of erythropoietin promoted recovery of hindlimb function, and the high dose of erythropoietin led to better outcome. High dose of erythropoietin exhibited a stronger suppressive effect on ferroptosis relative to the low dose of erythropoietin. The effects of erythropoietin on inhibiting ferroptosis-related protein expression and restoring mitochondrial morphology were similar to those of Fer-1 (a ferroptosis suppressor), and the effects of erythropoietin were largely diminished by RSL3 (ferroptosis activator). In vitro experiments showed that erythropoietin inhibited RSL3-induced ferroptosis in PC12 cells and increased the expression of xCT and Gpx4. This suggests that xCT and Gpx4 are involved in the neuroprotective effects of erythropoietin on spinal cord injury. Our findings reveal the underlying anti-ferroptosis role of erythropoietin and provide a potential therapeutic strategy for treating spinal cord injury.

Studies on the mechanism of Toxoplasma gondii Chinese 1 genotype Wh6 strain causing mice abnormal cognitive behavior.

BACKGROUND: Alzheimer's disease presents an abnormal cognitive behavior. TgCtwh6 is one of the predominant T. gondii strains prevalent in China. Although T. gondii type II strain infection can cause host cognitive behavioral abnormalities, we do not know whether TgCtwh6 could also cause host cognitive behavioral changes. So, in this study, we will focus on the effect of TgCtwh6 on mouse cognitive behavior and try in vivo and in vitro to explore the underlying mechanism by which TgCtwh6 give rise to mice cognitive behavior changes at the cellular and molecular level. METHODS: C57BL/6 mice were infected orally with TgCtwh6 cysts. From day 90 post-infection on, all mice were conducted through the open field test and then Morris water maze test to evaluate cognitive behavior. The morphology and number of cells in hippocampus were examined with hematoxylin-eosin (H&E) and Nissl staining; moreover, Aß protein in hippocampus was determined with immunohistochemistry and thioflavin S plaque staining. Synaptotagmin 1, apoptosis-related proteins, BACE1 and APP proteins and genes from hippocampus were assessed by western blotting or qRT-PCR. Hippocampal neuronal cell line or mouse microglial cell line was challenged with TgCtwh6 tachyzoites and then separately cultured in a well or co-cultured in a transwell device. The target proteins and genes were analyzed by immunofluorescence staining, western blotting and qRT-PCR. In addition, mouse microglial cell line polarization state and hippocampal neuronal cell line apoptosis were estimated using flow cytometry assay. RESULTS: The OFT and MWMT indicated that infected mice had cognitive behavioral impairments. The hippocampal tissue assay showed abnormal neuron morphology and a decreased number in infected mice. Moreover, pro-apoptotic proteins, as well as BACE1, APP and Aß proteins, increased in the infected mouse hippocampus. The experiments in vitro showed that pro-apoptotic proteins and p-NF-κBp65, NF-κBp65, BACE1, APP and Aß proteins or genes were significantly increased in the infected HT22. In addition, CD80, pro-inflammatory factors, notch, hes1 proteins and genes were enhanced in the infected BV2. Interestingly, not only the APP and pro-apoptotic proteins in HT22, but also the apoptosis rate of HT22 increased after the infected BV2 were co-cultured with the HT22 in a transwell device. CONCLUSIONS: Neuron apoptosis, Aß deposition and neuroinflammatory response involved with microglia polarization are the molecular and cellular mechanisms by which TgCtwh6 causes mouse cognitive behavioral abnormalities.

New insights into the evolutionary dynamic and lineage divergence of gasdermin E in metazoa.

Gasdermin (GSDM) is a family of pore-forming proteins that induce pyroptosis. To date, the origin and evolution of GSDM in Metazoa remain elusive. Here, we found that GSDM emerged early in Placozoa but is absent in a large number of invertebrates. In the lower vertebrate, fish, three types of GSDME, i.e., GSDMEa, GSDMEb, and a previously unreported type (designated GSDMEc), were idenitied. Evolutionarily, the three GSDMEs are distinctly separated: GSDMEa is closely related to tetrapod GSDME; GSDMEb exists exclusively in fish; GSDMEc forms the lineage root of tetrapod GSDMA/B/C/D. GSDMEc shares conserved genomic features with and is probably the prototype of GSDMA, which we found existing in all tetrapod classes. GSDMEc displays fast evolutionary dynamics, likely as a result of genomic transposition. A cross-metazoan analysis of GSDME revealed that GSDMEa shares a conserved caspase recognition motif with the GSDME of tetrapods and cnidarians, whereas GSDMEb has a unique caspase recognition motif similar to that of mammalian GSDMD, and GSDMEc exhibits no apparent caspase recognition motif. Through functional test, four highly conserved residues in vertebrate GSDME proved to be essential to auto-inhibition. Together our results provide new insights into the origin, evolution, and function of metazoan GSDMs.

Melatonin promotes sirtuin 1 expression and inhibits IRE1α-XBP1S-CHOP to reduce endoplasmic reticulum stress-mediated apoptosis in chondrocytes.

Osteoarthritis (OA) is the most common chronic disease characterized by a loss of chondrocytes and the degeneration of cartilage. Inflammation plays an important role in the pathogenesis and progression of OA via the activation of the endoplasmic reticulum (ER) stress signaling pathway. In this study, we stimulated human primary chondrocytes with lipopolysaccharide (LPS) to reduce cell viability and induce chondrocyte apoptosis. LPS-stimulated human primary chondrocytes induced ER stress and significantly upregulated the ER chaperone glucose-regulated protein 78 (GRP78) and increased the expression level of C/EBP-homologous protein (CHOP), a key mediator of ER stress--induced apoptosis. Interestingly, melatonin treatment attenuated ER stress-mediated chondrocyte apoptosis. Melatonin inhibited the expression of cleaved caspase-3, cleaved caspase-10, Bax, CHOP, GRP78, cleaved caspase-4, phospho-inositol-requiring enzyme 1α (P-IRE1α), and spliced X-box-binding protein 1 (XBP1S). In an anterior cruciate ligament transection mouse model of OA, melatonin (50 and 150 mg/kg) dose-dependently relieved joint cartilage degeneration and inhibitied of chondrocyte apoptosis. Immunohistochemical analysis indicated that melatonin could promote SIRT1 the expression and inhibit CHOP and cleaved caspase-3 expression in OA mice. In conclusion, our findings demonstrate for the first time that melatonin inhibits the IRE1α-XBP1S-CHOP signaling pathway by promoting the expression of SIRT1 in LPS-treated human chondrocytes and delaying OA progression in vivo.

3,3'-diindolylmethane inhibits LPS-induced human chondrocytes apoptosis and extracellular matrix degradation by activating PI3K-Akt-mTOR-mediated autophagy.

Osteoarthritis (OA) is a chronic degenerative joint disease characterized by articular cartilage destruction. The pathological mechanisms are complex; in particular, inflammation, autophagy, and apoptosis are often involved. 3,3-Diindolylmethane (DIM), a phytoconstituent extracted from cruciferous vegetables, has various effects such as anti-inflammatory, antioxidant and anti-apoptotic. However, the effects of DIM on osteoarthritic chondrocytes remain undetermined. In this study, we simulated a lipopolysaccharide (LPS)-induced osteoarthritis model in human primary chondrocytes. We found that LPS stimulation significantly inhibited autophagy, induced chondrocyte apoptosis and extracellular matrix (ECM) degradation, which could be ameliorated by DIM. DIM inhibited the expression of a disintegrin and metalloproteinase with thrombospondin motif 5 (ADAMTS-5), matrix metalloproteinase 13 (MMP13), cleaved caspase-3, Bax, and p62, and increased the expression level of collagen II, aggrecan, Bcl-2, light chain 3 Ⅱ (LC3 Ⅱ), and beclin-1. Mechanistic studies showed that DIM increased chondrocyte autophagy levels by inhibiting the activation of PI3K/AKT/mTOR pathway. In mice destabilization of the medial meniscus (DMM) model, immunohistochemical analysis showed that DIM inhibited the expression of p-PI3K and cleaved caspase-3, increased the expression of LC3 Ⅱ. Furthermore, DIM relieved joint cartilage degeneration. In conclusion, our findings demonstrate for the first time that DIM inhibits LPS-induced chondrocyte apoptosis and ECM degradation by regulating the PI3K/AKT/mTOR-autophagy axis and delays OA progression in vivo.

A Comparison of Mobile- and Fixed-Bearing Unicompartmental Knee Arthroplasties in the Treatment of Medial Knee Osteoarthritis: A Systematic Review and Meta-analysis of 1,861 Patients.

Many studies have compared mobile-bearing (MB) and fixed-bearing (FB) unicompartmental knee arthroplasties (UKAs) in patients with unicompartmental knee osteoarthritis (OA). The present systematic review and meta-analysis examined the differences in the clinical and radiological outcomes of MB UKA and FB UKA. PubMed, EMBASE, and Cochrane databases, as well as Google Scholar were searched for relevant studies. Randomized controlled trials (RCTs) and cohort studies that compared MB UKA and FB UKA were included. The weighted mean difference in the knee scores and range of motion (ROM) as well as the summary odds ratio of postoperative mechanical axis alignment, radiolucency, revision rate, and complications were calculated in the MB UKA and FB UKA groups. Finally, 2 RCTs and 11 cohort studies that involved 1,861 patients (1,996 knees) were included. The FB UKA group showed better postoperative Knee Society score (KSS), Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), and ROM than the MB UKA group. However, the MB UKA group had more knees with a neutral limb alignment and a lower incidence of polyethylene wear than the FB UKA group. No significant differences were observed between the groups with respect to radiolucency, revision rate, and complications, such as arthritis progression, aseptic loosening, and postoperative pain. This meta-analysis has demonstrated that both prostheses provided excellent clinical outcomes and survivorship in patients with unicompartmental knee OA. The MB UKA group achieved the expected postoperative neutral limb alignment as compared with the FB UKA group, while the FB UKA group showed higher knee scores and superior ROM than the MB UKA group. Limited evidence is currently available; therefore, the results of our meta-analysis should be interpreted with caution.

Effect of fucoidan on ethanol-induced liver injury and steatosis in mice and the underlying mechanism.

BACKGROUND: Alcoholic liver disease is caused as a result of chronic alcohol consumption. In this study, we used an alcoholic liver injury mouse model to investigate the effect of fucoidan on ethanol-induced liver injury and steatosis and the underlying mechanisms. METHODS: All mice were randomly divided into four groups: 1) control group, 2) model group, 3) diammonium glycyrrhizinate treatment group (200 mg/kg body weight), and 4) fucoidan treatment group (300 mg/kg body weight). Administration of ethanol for 8 weeks induced liver injury and steatosis in mice. RESULTS: Fucoidan treatment decreased serum alanine aminotransferase activity, serum total cholesterol levels, and hepatic triglyceride levels, and improved the morphology of hepatic cells. Fucoidan treatment upregulated the expression of AMPKα1, SIRT1, and PGC-1α and inhibited the expression of ChREBP and HNF-1α. The levels of hepatic IL-6 and IL-18 were significantly decreased in the fucoidan group. Further, the levels of cytochrome P450-2E1 (CYP2E1), glucose-regulated protein (GRP) 78, and 3-nitrotyrosine (3-NT) in hepatic tissues were reduced in the fucoidan group as compared to the model group. Fucoidan significantly reversed the reduction of ileac Farnesoid X receptor (FXR) and fibroblast growth factor 15 (FGF15) levels induced by alcohol-feeding and reduced CYP7A1 (cholesterol 7α-hydroxylase) expression and total bile acid levels in the liver tissue. In addition, fucoidan regulated the structure of gut flora, with increased abundance of Prevotella and decreased abundance of Paraprevotella and Romboutsia as detected by 16S rDNA high-throughput sequencing. CONCLUSION: Fucoidan inhibited alcohol-induced steatosis and disorders of bile acid metabolism in mice through the AMPKα1/SIRT1 pathway and the gut microbiota-bile acid-liver axis and protected against alcohol-induced liver injury in vivo.

The New Role of Sirtuin1 in Human Osteoarthritis Chondrocytes by Regulating Autophagy.

OBJECTIVE: The aim of this study is to investigate the role of Sirtuin1 (Sirt1) in the regulation of autophagy for human osteoarthritis (OA) chondrocytes. DESIGN: All cartilage samples were collected from human donors, including young group, aged group, and OA group. Primary chondrocytes were isolated and cultured with Sirt1 activator or inhibitor. Sirt1 expression in cartilage tissue and chondrocytes was evaluated, and the deacetylation activity of Sirt1 was determined. The alteration of autophagy activity after upregulating or downregulating Sirt1 was detected. Chondrocytes were treated with autophagy activator and inhibitor, and then the protein level of Sirt1 was examined. The interactions between Sirt1 and autophagy-related proteins Atg7, microtubule associated protein 1 light chain 3 (LC3), and Beclin-1 were determined by using immunoprecipitation. RESULTS: The assay of articular cartilage revealed that the expression of Sirt1 might be age-related: highly expressed in of younger people, and respectively decreased in the elderly people and OA patients. In vitro study was also validated this result. Further study confirmed that higher levels of Sirt1 significantly increased autophagy in aged chondrocytes, while the lower expression of Sirt1 reduced autophagy in young chondrocytes. Of note, the high levels of Sirt1 reduced autophagy in OA chondrocytes. When the chondrocytes were treated with autophagy activator or inhibitor, we found the expression of Sirt1 was not affected. In addition, we found that Sirt1 could interact with Atg7. CONCLUSION: These results suggest that Sirt1 in human chondrocytes regulates autophagy by interacting with autophagy related Atg7, and Sirt1 may become a more important target in OA treatment.

[Complication after selective arterial embolization in internal iliac artery and median sacral artery with gelfoam particle in dogs].

OBJECTIVE: To observe the complication after embolizing the bilateral internal iliac arteries and the median sacral artery of dogs by different combinations and embolization levels with gelfoam particle, and to provide a reference for safety application of gelfoam in clinic. METHODS: Sixteen common grade adult healthy dogs (weighing 10-13 kg, 14 males and 2 females) were randomly divided into 5 groups. Under the monitoring of digital subtraction angiography (DSA), the embolization was performed with gelfoam particle (diameter, 50-150 microm) in bilateral internal iliac arteries and the main branch of the median sacral artery (group A, n = 3), in bilateral internal iliac arteries and the first branch of the median sacral artery (group B, n = 3), in the main branch of bilateral internal iliac arteries (group C, n = 3), in the unilateral internal iliac artery and the main branch of the median sacral artery (group D, n = 4), and in the main branch of unilateral internal iliac artery (group E, n = 3). Under the DSA, the anatomic relationships of the abdominal aorta, bilateral external iliac arteries, bilateral internal iliac arteries, and median sacral artery were observed before embolization. The survival dogs were observed and the specimens of bladder, rectum, sciatic nerve, and gluteal muscles were harvested for the general and histological observations at 3 days after embolization. RESULTS: In dogs, there was no common iliac artery; bilateral external iliac arteries originated from the abdominal aorta and the starting of the median sacral artery had variation. Seven dogs (3 in group A, 3 in group C, and 1 in group D) died within 2 days after embolization, and the others survived to the end of the experiment. In the dead dogs of groups A, C, and D, the darkening and necrosis of the rectum were observed; the bladder presented lamellar obfuscation and focal hemorrhage and edema; and the median urinary volume in bladder was 270.6 mL. In survival dogs, no obvious change was observed in the rectum; the bladder only manifested light edema; and the median urinary volume in bladder was 137.0, 220.5, and 28.0 mL, respectively in groups B, D, and E. The rectum and bladder of dead dogs in groups A, C, and D manifested the disrupted cells, generous inflammatory cells infiltration, and desquamation of epithelial cells; the rectum and bladder of survival dogs in groups B, D, and E manifested light inflammatory cells infiltration and edema; the embolized artery mainly focused on the arterioles whose diameter was 100-200 microm. The sciatic nerve and gluteal muscles of each group had no obvious change except for light edema. CONCLUSION: When the internal iliac artery and median sacral artery are embolized with gelfoam particle with a diameter of 50-150 microm, to ensure the safeness of pelvic organs, the embolized artery can not exceed the first branch when the 3 arteries are embolized at the same time, or reserve at least unilateral internal iliac artery when embolized to the trunk, or it will result in pelvic organ necrosis and perforation.