Expression Levels Of Fas Research Articles

Cadmium impairs the development of natural killer cells and bidirectionally modifies their capacity for cytotoxicity

Cadmium (Cd) is a highly toxic heavy metal in the environment. The aim of this study was to investigate the impact of Cd on natural killer (NK) cells. C57BL/6 mice were treated with 10 ppm Cd via drinking water for 3 months, and the development of NK cells in the bone marrow (BM) and the cytotoxicity of mature NK (mNK) cells in the peripheral immune organs were evaluated thereafter; the impact of Cd on the cytotoxicity of mNK cells from human peripheral blood mononuclear cells (PBMC) was also investigated. Whereas Cd treatment impaired the differentiation of NK progenitors in the BM, Cd treatment activated the JAK3/STAT5 signaling to drive the proliferation of mNK cells and thereby lead to a compensation increase of mNK cells in the peripheral immune organs of mice. Additionally, Cd treatment bidirectionally regulated the cytotoxicity of mouse mNK cells to differential tumor cells, dependent on the levels of Fas expression in the tumor cells; mechanically, Cd treatment activated the JAK3/STAT5 signaling to promote the expression of FasL in mNK cells to increase their cytotoxicity, while Cd treatment reduced the expression of granzyme B in mNK cells to impair their cytotoxicity in the peripheral immune organs of mice. Likewise, in vitro assays indicated that Cd treatment also activated the JAK3/STAT5 signaling to increase the expression of FasL, whereas Cd treatment reduced the expression of granzyme B in human mNK cells. Thus Cd treatment impaired the development of NK cells in the BM and bidirectionally regulated the cytotoxicity of mNK cells in the peripheral immune organs, which may extend our current understanding for the immunotoxicity of Cd.

A soy glycinin derived octapeptide protects against MCD diet induced non-alcoholic fatty liver disease in mice

Soy glycinin derived octapeptide (SGP8) is a peptide obtained from degradation of the soy glycinin, whose amino acid sequence is IAVPGEVA. To determine the effect of SGP8 on non-alcoholic fatty liver disease (NAFLD), steatosis HepG2 cells were induced by 1 mmol/L free fatty acid (FFA) and C57BL/6J mice were fed with methionine-choline deficient (MCD) diet for 3 weeks to establish NAFLD model. The results of oil red O staining and total cholesterol (TC)/triglyceride (TG) contents showed that SGP8 could significantly reduce the lipid content of steatosis HepG2 cells. In vivo, SGP8 lowered plasma alanine aminotransferase (ALT) and low density lipoprotein (LDL) content, normalized hepatic superoxide dismutase (SOD) and malondialdehyde (MDA) production, and reduced the severity of liver inflammation. The results of Western blotting showed that SGP8 increased expression of Sirtuin-1 (SIRT1) and phosphorylation level of AMP activated protein kinase (AMPK) in hepatocytes. Through activation of SIRT1/AMPK pathway, SGP8 downregulated the expression of sterol regulatory element binding protein 1c (SREBP-1c) and its target genes ACC and FAS expression levels, and increased the phosphorylation level of acetyl CoA carboxylase (ACC). Furthermore, SGP8 also upregulated the expression of transcription factor peroxisome proliferator activated receptor α (PPARα), which was regulated by SIRT1/AMPK pathway, and its target gene CPT1 level. In conclusion, SGP8 might improve NAFLD by activating the SIRT1/AMPK pathway. Our data suggest that SGP8 may act as a novel and potent therapeutic agent against NAFLD.

Abstract 2825: Stromal cell derived IL6 inhibits the extrinsic apoptotic pathway in multiple myeloma cell lines

Abstract Multiple myeloma (MM) is a disease of malignant plasma cells that resides in the bone marrow microenvironment (BMM). While advances in therapy have improved patient outcomes, the majority of patients relapse. Clinical trials have shown that BCMA targeted CAR-T therapy induces MM remission in most patients with heavily pretreated relapsed refractory MM; however, the duration of the response has been disappointing. CAR-T cells induce target cell death through two primary mechanisms, granzyme B and FASL. To define the role of FAS-induced cell death in CAR-T therapy in MM, we first evaluated if FAS-induced cell death was important for CAR-T activity in MM. We utilized CRISPR-Cas9 to generate FAS KOs in three MM cell lines: RPMI8226, KMS18, and OCIMY5. These lines were utilized in vitro in cytotoxic CAR-T assays where MM cells were cultured with BMCA CAR-T cells at five effector to target (E:T) ratios (range: 0:1 to 1:1). The loss of FAS protected KMS18 and OCIMY5 from CAR-T induced cell death, changing the E:T ratio needed to induce 50% MM cell death 3.2 and 3.4 fold respectively, but did not protect RPMI8226. We determined the expression of FAS negatively correlated with the EC50 of rFASL, suggesting that FAS expression levels are important for this response. To determine the role of the intrinsic apoptotic pathway in FASL-induced death, we knocked out BAK and BAX in KMS12 PE and OCIMY5. This did not protect KMS12 PE or OCIMY5 from rFASL-induced cell death, suggesting that FAS uses the type I mitochondria-independent pathway in these cell lines. The BMM supports MM progression and promotes drug resistance in part by altering the apoptotic threshold. Therefore, we examined the impact of the BMM on FASL-induced cell death. Coculture of MM cells with the HS5 stromal cell line protected KSM12 PE and KMS18 from rFASL, with the EC50 of rFASL changing 2.72 and 3.1 fold respectively, but did not protect OCIMY5. To determine if the protection observed was cell contact dependent, or only required soluble factors, we utilized HS5 conditioned media (CM). HS5 CM protected KMS12 PE, KMS18, and surprisingly OCIMY5 from rFASL, causing a change in EC50 from control of 1.65 - 2.23 fold. CM lacking IL6 did not protect MM cells from rFASL. Therefore, we tested the effects of IL6 addition IL6(10 ng/ml) and observed protection from rFASL in KMS12 PE and KMS18 cells with a change in EC50 of 2.03 and 3.84 fold respectively; however, we did not see protection in OCIMY5. To identify the mechanism of IL6-induced resistance to rFASL, we cultured BAK/BAX KO cells in HS5 CM and observed protection of these apoptosis-deficient KMS12 PE and OCIMY5 cells from rFASL. This suggests that soluble factors in the BMM can convert cells from type I to type II (mitochondrial-dependent) death receptor signaling and that inhibiting BMM signals such as IL6 may enhance FASL-induced cell death in MM. However, the data also demonstrate that the role of FAS in CAR-T killing is cell context dependent. Citation Format: James C. Ackley, Samuel McCachren, Sagar Lonial, Damien J. Green, Stanley R. Riddell, Geoffrey R. Hill, Madhav V. Dhodapkar, Lawrence H. Boise. Stromal cell derived IL6 inhibits the extrinsic apoptotic pathway in multiple myeloma cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2825.

Amycenone reduces excess body weight and attenuates hyperlipidaemia by inhibiting lipogenesis and promoting lipolysis and fatty acid β-oxidation in KK-Ay obese diabetic mice.

Excess body weight and hyperlipidaemia cause severe health problems and have social implications. Amycenone is an active substance extracted from Yamabushitake mushrooms with no reports of its activity against excess body weight and hyperlipidaemia. This research clarifies the effects and mechanisms of action of amycenone on the inhibition of body weight excess and hyperlipidaemia attenuation using KK-Ay mice. Amycenone or water was administered to 8-week-old male KK-Ay mice by gavage for 8 weeks. Their body weight and food intake were recorded during the experiment. At the end of the experimental period, the mice were dissected, and blood samples, lipid metabolism-related organs and tissues were collected and stored for further analysis. Amycenone treatment suppressed body weight gain and improved serum levels of fasting blood glucose and non-esterified fatty acids. Additionally, serum and hepatic cholesterol and triacylglycerol levels were reduced after this treatment, whereas the phosphorylation levels of AMPK, PKA and HSL increased and the expression level of FAS decreased. The protein level of C/EBPβ and gene expression level of Cpt1 were higher in the perirenal adipose tissue of amycenone-treated KK-Ay mice. Furthermore, amycenone phosphorylated AMPK, PKA and ACC, and PPARγ expression was lower in the mesenteric adipose tissue. The phosphorylation levels of AMPK, LKB1, PKA and ACC were also induced, and FAS expression level was reduced in the liver of the amycenone-treated group. Amycenone could reduce excess body weight and attenuate hyperlipidaemia in KK-Ay mice by inhibiting lipogenesis and promoting lipolysis through lipid metabolism pathway stimulation and fatty acid β-oxidation acceleration.

Matrine inhibits ovarian cancer cell viability and promotes apoptosis by regulating the ERK/JNK signaling pathway via p38MAPK.

Ovarian cancer displays the highest mortality rate among all types of gynecological cancer worldwide. The survival of patients with ovarian cancer remains poor due to poor responses to anticancer treatments. The present study aimed to investigate the therapeutic effects and potential mechanism underlying matrine in ovarian cancer tissues, ovarian cancer cells and a CAOV-3-derived tumor-bearing mouse model. MTT, migration, invasion, flow cytometry, immunofluorescence and immunohistochemistry assays were performed to assess the inhibitory effects of matrine on ovarian cancer. A xenograft ovarian cancer mouse model was established and treated with matrine or PBS. The results demonstrated that compared with the control group, matrine significantly induced ovarian cancer cell apoptosis by upregulating caspase-8 and Fas cell surface death receptor (Fas) expression levels, and downregulating Bcl-2 and Bcl-xl expression levels. Moreover, compared with the control group, matrine significantly inhibited ovarian cancer cell viability, migration and invasion by downregulating metastasis associated protein-1, fibronectin, angiotensin II type 2 receptor–interacting protein 3a and H high mobility group AT-hook 2 expression levels. Compared with the control group, matrine significantly increased p38MAPK, phosphorylated (p)ERK/ERK and pJNK/JNK expression levels in ovarian cancer cells. p38MAPK knockdown significantly downregulated p38MAPK, pERK/ERK and pJNK/JNK expression levels compared with the control group, which significantly promoted ovarian cancer cell viability, migration and invasion. In vivo experiments demonstrated that compared with the control group, matrine significantly suppressed tumor growth by markedly upregulating p38MAPK, ERK and JNK expression levels. The immunohistochemistry results demonstrated that caspase-8 and Fas expression levels were notably increased, whereas Bcl-2 and Bcl-xl expression levels were obviously decreased in matrine-treated tumors compared with PBS-treated tumors. In conclusion, the present study demonstrated that matrine inhibited ovarian cancer cell viability, migration and invasion, but induced cell apoptosis, suggesting that matrine may serve as a promising anticancer agent for the treatment of ovarian cancer.

Co-culture model reveals the characteristics of theca cells and the effect of granulosa cells on theca cells at different stages of follicular development.

Theca cells (TCs) play an important role in follicular development, which cannot be separated from granulosa cells (GCs). However, compared with mammals, the TCs and the effects of GCs on TCs at different follicular development stages (FDSs) have specific characteristics in avian species, but none of them have been clearly defined. In this study, we established an in vitro co-culture (with GC at the corresponding stage) model of goose TCs at different FDSs (pre-hierarchical, hierarchical and F1) by using a transwell system. The properties of TCs in co-culture at the three FDSs, including cell morphology, activity and intracellular lipid content, as well as the expression of key genes involved in de novo lipogenesis, steroidogenesis, proliferation and apoptosis, were examined and defined. We further compared the mono-culture and co-culture groups. After co-culture, the activity of TCs showed significant (p<.01) increases in all stages; moreover, in pre-hierarchical TCs, the expression levels of FAS, SREBP, 3β-HSD and CCND1 were promoted, and PPARγ, CYP19, BCL2 and CAS3 were inhibited (p<.05); in the hierarchical TCs, the expression levels of PPARγ, FAS, CYP19, CCND1 and BCL2 were promoted, and SREBP, STAR, 3β-HSD and CAS3 were inhibited (p<.05), whereas in the F1 TCs, the expression levels of PPARγ, FAS, 3β-HSD, CYP19 and CCND1 were promoted, and STAR and CAS3 were inhibited (p<.05). These results suggested that GCs at the three FDSs have dynamic and complex influences on the physiological characteristics of TCs, and the influences on TCs at the three FDSs were varied.

Dietary DHA/EPA ratio affects growth, tissue fatty acid profiles and expression of genes involved in lipid metabolism in mud crab Scylla paramamosain supplied with appropriate n-3 LC-PUFA at two lipid levels

An 8-week feeding trial was conducted to determine the optimal dietary docosahexaenoic acid/eisosapentaenoic acid (DHA/EPA) ratio of mud crab (Scylla paramamosain) supplied with optimal n-3 LC-PUFA at two dietary lipid levels. Eight isonitrogenous diets were formulated to contain 7% and 12% crude lipid, each with DHA/EPA ratios of 0.6, 1.2, 2.3 and 3.2, respectively. Each diet was randomly assigned to triplicate groups of 30 juvenile mud crabs (initial weight 20.9 ± 0.6 g) that were stocked in single crab cells. In crabs fed 7% lipid, the diet with a DHA/EPA ratio of 2.3 showed significantly higher weight gain than crabs fed the other ratios while in crabs fed 12% lipid, lower weight gain and specific growth rate were observed in crabs fed the diet with a DHA/EPA ratio of 0.6 than crabs fed the other ratios. Lipid content in hepatopancreas significantly increased as dietary DHA/EPA ratio increased from 1.2 to 2.3 in crabs fed 7% lipid, while no differences were observed among crabs fed the diets with DHA/EPA ratios higher than 0.6 when fed 12% lipid. Total fatty acid and DHA contents and DHA/EPA ratio showed increasing, and EPA decreasing, trends in muscle and hepatopancreas with increased dietary DHA/EPA ratio, at both dietary lipid levels. The hemolymph triacylglycerol and total cholesterol contents were higher in crabs fed dietary DHA/PA ratios of 1.2 and 2.3 than those fed ratios of 0.6 and 3.2 at 7% dietary lipid, and lowest low and high-density lipoprotein cholesterol contents were observed in crabs fed DHA/EPA dietary ratios of 0.6 and 3.2 at 7% and 12% lipid, respectively. The expression levels of fas, aco3 and fatp4 were significantly up-regulated, and cptI, hsl and ldlr were down-regulated, with increased dietary DHA/EPA ratio in crabs fed 7% lipid. In crabs fed 12% lipid, the expression levels of g6pd, 6pgd, srebp-1, aco1 and fatp4 were down-regulated, and fabp-1 was up-regulated, with increased dietary DHA/EPA ratio. The expression levels of elovl4 and Δ6 fad initially increased and then decreased as dietary DHA/EPA ratio increased from 0.6 to 3.2 in crabs fed both 7% and 12% lipid. Based on analysis of weight gain versus dietary DHA/EPA ratio, the optimal dietary DHA/EPA ratios of mud crab S. paramamosa were estimated to be 2.2 and 1.2 when supplied with optimal n-3 LC-PUFA at 7% and 12% lipid, respectively.

Effects of different dietary oil sources on growth performance, antioxidant capacity and lipid deposition of juvenile golden pompano Trachinotus ovatus

Vegetable oils (VO) that are used to substitute fish oil in aquafeeds may affect, not only the fatty acid composition, but also lipid metabolism and distribution. The present study was designed to investigate this issue in juvenile golden pompano Trachinotus ovatus fed eight diets formulated with typical VO with widely varying fatty acid compositions including coconut oil (CO), palm oil (PO), oil-tea camellia seed oil (OTO), olive oil (OO), canola oil (CNO), peanut oil (PNO), linseed oil (LO) and perilla oil (PFO), in comparison with fish fed fish oil (FO). After the 8-week feeding trial, fish fed the CO diet had the highest growth performance, and higher general antioxidant capacities in serum and liver than in fish fed the other VO. The crude lipid content in whole body and expression levels of fas were lower in fish fed the FO, PFO and LO diets, while lipid contents and expression levels of scd were higher in fish fed the OTO and PNO diets. Other than fish fed the PFO diet, the total lipid contents of liver in other fish fed the other VO diets were higher than that in fish fed the FO diet, with the highest contents in fish fed the OTO and OO diets. The expression levels of genes involved in fatty acid catabolism and transport, namely pparα, cpt1 and apoB100, were higher in fish fed diet PFO than in fish fed the other diets. Comparing the fatty acid compositions of tissues and diets showed that 18:1n-9, 18:3n-3 (ALA) and 22:6n-3 (DHA) were preferentially deposited in tissues of pompano, with DHA preferentially deposited in polar lipids rather than neutral lipids. However, excessive dietary ALA in PFO did not lead to increased deposition of ALA, but increased liver lipid content. The present study showed that dietary lipid sources had significant influences on growth performance and antioxidant capacity, as well as on lipid deposition. Low dietary 18:1n-9, high n-3 long-chain polyunsaturated fatty acids and an appropriate ratio of ALA/LNA (18,2n-6) could reduce lipid deposition in pompano tissues, especially liver.

Danggui Niantong decoction induces apoptosis by activating Fas/caspase-8 pathway in rheumatoid arthritis fibroblast-like synoviocytes

To explore the effect of Danggui Niantong decoction (DGNTD) on cell apoptosis and TNF receptor super family 6 (Fas)/caspase-8 pathway in rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLS). FLS isolated from the synovial tissue of RA patients were cultured and identified using immunofluorescence staining. The cells were treated with 10% blank serum (blank control group), 10% sera containing low, moderate or high doses of DGNTD, or 20 μmol/mL KR-33493 (a Fas inhibitor) combined with 10% serum containing high-dose DGNTD. MTT assay was used to detect the proliferation of the cells after the treatments. Apoptosis of the cells was detected at 48 h in each group using Hoechst 33342 staining and flow cytometry with annexin V-FITC/PI staining. The mRNA and protein expressions of Fas, FADD, caspase-8 and caspase-3 in the cells at 48 h were detected using qPCR and Western blotting. Immunofluorescence staining identified the cultured cells as FLS. Treatment with DGNTD-containing sera significantly inhibited the proliferation of FLS, and the inhibitory effects were enhanced as the dose and intervention time increased (P &lt; 0.05). Hoechst 33342 staining and flow cytometry showed that the sera containing different doses of DGNTD significantly promoted apoptosis of FLS (P &lt; 0.05). The expression levels of Fas, FADD, caspase-8, and caspase-3 at both mRNA and protein levels were significantly increased in the cells after treatment with different doses of DGNTD-containing sera (P &lt; 0.05). The application of KR-33493 obviously reversed the effects of DGNTD on the FLS (P &lt; 0.05). DGNTD can induce apoptosis of the FLS by activating Fas/caspase-8 signaling pathway.

Potential use of C-phycocyanin in non-alcoholic fatty liver disease

C-phycocyanin (C-PC) is a kind of photosynthetically assisted pigment, which is ubiquitous in cyanobacteria cells. We investigated the effect of C-PC on non-alcoholic fatty liver disease (NAFLD) and its mechanism. Through oil red O staining, TC/TG detection, liver SOD/MDA detection and liver H&E staining, we found that C-PC could significantly reduce the lipid accumulation in the steatosis L02 cells and the liver of non-alcoholic steatohepatitis (NASH) mice, and improve the antioxidant capacity of liver. The results of Western Blotting showed that C-PC upregulated the expression of AMPK phosphorylation and downregulated SREBP-1c and its target genes ACC and FAS expression levels. Furthermore, C-PC also upregulated the expression of transcription factor PPARα, which was regulated by AMPK, and its target genes CPT1 level. In addition, C-PC could promote AMPK phosphorylation in hepatocytes while increasing the phosphorylation level of ACC in vivo and in vitro. Besides, C-PC could also improve the liver inflammatory infiltration by upregulated the expression of PPARγ and downregulated the expression of CD36, IL6 and TNFα. These results indicate that C-PC may improve hepatic lipid accumulation and inflammation in the non-alcoholic fatty liver mice by activating AMPK pathway of hepatocytes. The finding provides important help for the research and development of C-PC in the nutraceuticals and therapeutics of NAFLD.

Effects of Lactobacillus Casei YBJ02 on Lipid Metabolism in Hyperlipidemic Mice.

Traditionally fermented yak yogurt as a Tibetan dairy product is high not only in nutrients but also in probiotics. A probiotic strain with a potential lipid reducing effect was isolated from yak yogurt. An animal model for hyperlipidemia was evaluated using the blood index and expression levels of lipid metabolism-related proteins in mice to determine the effect of Lactobacillus casei YBJ02 (LC-YBJ02) on lipid metabolism in hyperlipidemic mice and the underlying mechanism. LC-YBJ02 at different concentrations exhibited certain inhibitory effects on the increase in blood lipid in mice. Particularly, high concentrations of LC-YBJ02 can reduce the cholesterol, triglycerides, and low-density lipoprotein content; however, no significant effect on the high-density lipoprotein of the body has been reported. LC-YBJ02 can effectively increase the reduction in cholesterol level by fecal excretion. In this study, the gene-affecting mouse obesity was determined using experimental results in mice. The expression levels of peroxisome proliferator-activated receptor γ (PPARγ), CCAAT enhancer binding protein alpha (CEBP)/α, sterol regulatory element binding protein (SREBP)-1c, and FAS could improve because of the high-fat diet in kidney fat. Bacteria at different concentrations could also decrease these expression levels. Specifically, the high concentration of LC-YBJ02 could suppress the expression of PPARγ, CEBP/α, and SREBP-1c; however, the expression of FAS was not significantly inhibited. PPARγ and FAS expression levels in the liver were low, but no significant difference was indicated. CEBP/α and SREBP-1c expression in mouse liver was further detected by Western blot analysis; CEBP/α was considerably low and could not be detected. On the basis of these results, LC-YBJ02 could be used as probiotics through its lipid reduction effects.

Gadolinium chloride alleviates partial warm ischemia-reperfusion injury of liver by suppressing kupffer cell function in mice

Objective To investigate the effect and mechanism of gadolinium chloride alleviates partial warm ischemia-reperfusion injury of liver by suppressing kupffer cell function in mice. Methods Forty-eight male C57BL/6 mice were randomly divided into sham-operated group (Sham), Hepatic ischemia-reperfusion injury (HIRI) group, gdolinium chloride groups (Gd+ IRI), and cobalt protoporphyrin (CoPP+ IRI) group (n=12). The rats were injected with the same dose of normal saline in Sham group and IRI group. Gd+ IRI group were intraperitoneally injected with GdCl3 (20 mg/kg) 24 hours before operation. At the same time, CoPP+ IRI group were intraperitoneally injected with CoPP (5 mg/kg). Except for Sham group, an 70% volume HIRI model was established by means of 60 minutes ischemia and then 6 hours reperfusion in the other groups. The levels of serum aspartate transaminase (AST) and alanine aminotransferase (ALT) in each group were compared. Enzyme-linked immunosorbent assay were used to test the levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and malondialdehyde (MDA) in the liver tissue. The level of tumor necrosis factor-α (TNF-α), interleukin (IL)-1β, IL-6, heme oxygenase-1 (HO-1), and protective factor IL-10 mRNA level were detected by real time quantitative polymerase Chain Reaction (RT-PCR). Immunohistochemical staining was used to measure Fas, FasL protein expression level of extrahepatic biliary epithelial cells. Results Respectively, the serum ALT level in the HIRI, Gd+ IRI and CoPP+ IRI groups was (1 236.7±62.4), (638.9±50.9), (740.3±53.1) U/L, and the AST level was (1 107.8±68.3), (568.5±42.7), (679.1±49.9) U/L, the serum level of ALT and AST in Gd+ IRI and CoPP+ IRI groups was lower than IRI group (t=6.721, P<0.01; t=4.732, P<0.05; t=5.984, P<0.01; t=2.691, P<0.05). Compared with IRI group, the activity of SOD and GSH-Px was promoted (t=7.301, P<0.01; t=1.937, P<0.05), whereas the levels of MDA was reduced (t=9.519, P<0.01; t=8.635, P<0.01). Simultaneously, the TNF-α, IL-1β and IL-6 mRNA level in Gd+ IRI and CoPP+ IRI groups were decreased (t=6.721, P<0.01; t=2.316, P<0.05), and the levels of IL-10 and HO-1 mRNA were increased (t=2.973, P<0.05; t=7.921, P<0.01). After hepatic artery reflow 6 h, the Fas expression levels on bile ducts epithelial cells in Gd+ IRI and CoPP+ IRI groups respectively was 22.4%, 29.7%, which was lower than IRI group (38.9%) (χ2=6.561, P<0.01; χ2=2.547, P<0.05). The FasL expression levels in Gd+ IRI and CoPP+ IRI groups respectively was 21.2%, 25.1%, which was lower than IRI group (36.5%) (χ2=5.982, P<0.05; χ2=3.188, P<0.05). Conclusion Gdolinium chloride shows an protective function aganist liver ischemia-reperfusion iniury in mice by Oxidative stress, inhibiting inflammation and cell apoptosis. The up-regulation of HO-1 by gdolinium chloride may be the possible mechanism. The up-regulation of HO-1 and inhibiting the Fas/FasL protein expression by gdolinium chloride may be the possible mechanism. Key words: Gdolinium chloride; Liver; Ischemia-reperfusion injury; Heme oxygenase-l

Synergistic effect of acupuncture and moxibustion on expression of renal Fas and FasL mRNAs and proteins in contrast-induced nephropathy diabetic rats

To observe the effect of acupuncture and moxibustion on renal Fas and Fas ligand (FasL) expression in contrast-induced nephropathy (CIN) diabetic rats, so as to explore its protective mechanisms. A total of 40 diabetic male SD rats were randomly divided into control, model, acupuncture, moxibustion and combined treatment (acupuncture combined with moxibustion) groups (n＝8 rats in each group). The diabetic model was established by i．p. injection of Streptozotocin (60 mg/kg), followed by regular raising for 9 weeks. Then, the manual acupuncture, moxibustion or acupuncture plus moxibustion at "Sanyinjiao"(SP6) "Shenshu"(BL23) and "Pishu"(BL20) was given to the diabetic rats, once daily for 7 days. On the 8th day, these diabetic rats received i．p. injection of compound Meglumine Diatrizoate Injection (10 mL/kg) for establishing CIN model. Then, the venous blood specimens were collected for detecting contents of serum urine nitrogen (BUN, with urease method) and creatinine (Scr, with picric acid method). The kindey tissues were exteriorized for detecting nitricoxide synthase (NOS, with oxidation-reduction method), malondialdehyde (MDA, with thiobarbituric acid method), superoxide dismutase (SOD, with xanthine oxidase technique) and total antioxidant capacity (T-AOC, with colorimetric technique). The expression levels of Fas and FasL in the kidney tissues were detected by quantitative real-time PCR and Western blot separately. The histopathological and ultrastructural changes of the kidney were observed under microscope (after H．E. staining) and transmission electron microscope, respectively. Following modeling and compared with the control group, the BUN, Scr and renal MDA contents, and Fas and FasL mRNA and protein expression levels were significantly increased (P<0.01, P<0.05), and renal NOS, SOD and T-AOC levels considerably decreased in the model group (P<0.05). After the interventions and compared with the model group, serum BUN and Scr contents and renal Fas mRNA and protein expression levels in both acupuncture group and combined treatment group, BUN in the moxibustion group, and renal MDA content and FasL mRNA and protein expression levels in the 3 treatment groups were obviously down-regulated (P<0.05，P<0.01), while renal NOS in the 3 treatment groups, and SOD and T-AOC levels in the combined treatment group were considerably up-regulated (P<0.01，P<0.05). The effects of the combined treatment were significantly superior to those of simple acupuncure and simple moxibustion in down-regulating serum BUN contents, and Fas and FasL mRNA and protein expression levels, and in up-regulating renal NOS, SOD and T-AOC activity (P<0.05). No significant differences were found between the acupuncture and moxibustion groups in down-regulating serum BUN, Scr and renal MDA contents, and Fas and FasL mRNA and protein levels, as well as in up-regulating NOS content (P>0.05). H．E. staining showed lobular changes of glomeruli, many local necrosis and vascular degeneration in the model group, and transmission electron microscope displayed thickened basement membrane of the glomerular blood loop, many vacuoles in the cytoplasm of renal tubular epithelial cells, severely swollen mitochondria with disordered, broken or disappeared cristae, and some apoptotic cells in the model group, which were relatively milder in the combined treatment group. Joint application of acupuncture and moxibustion can reduce the oxidative stress and renal injury in CIN diabetic rats, which may be associated with its function in down-regulating the expression of renal Fas and FasL genes and proteins. Acupuncture and moxibustion has a synergistic effect.

Effect of divergence in residual feed intake on expression of lipid metabolism-related genes in the liver of meat-type ducks1

Lipid metabolism is considered one of the important factors affecting residual feed intake (RFI). However, the relationship between RFI and expression of lipid metabolism-related genes is unknown in meat-type ducks. To address this issue, a total of 1,000 male meat-type ducks with similar body weight were randomly selected to measure body weight gain and feed intake from 21 to 42 d of age to estimate RFI. The 8 greatest- (high RFI [HRFI]) and lowest- (low RFI [LRFI]) ranking birds were then selected for the present study. Relative expressions of key genes, namely sirtuin 1 (Sirt1), forkhead box O1 (Foxo1), peroxisome proliferator-activated receptor gamma (PPARγ), sterol regulatory element-binding transcription factor 1c (SREBP-1c), fas cell surface death receptor (FAS), acetyl-CoA carboxylase alpha (ACC), carnitine palmitoyltransferase 1A (CPT1A), and acyl-CoA oxidase 1 (ACOX1), were then determined in the HRFI and LRFI ducks by quantitative PCR. The results showed that RFI, feed conversion ratio (FCR), and average daily feed intake (ADFI) were significantly lower (P < 0.05) in LRFI ducks than in HRFI ducks. In addition, expression of Sirt1, Foxo1, CPT1A, and ACOX1 were significantly higher in LRFI ducks than in HRFI ducks (P < 0.05), whereas PPARγ and FAS expression levels were significantly lower in LRFI ducks than in HRFI ducks (P < 0.01). Correlation analysis showed that Sirt1, CPT1A, and ACOX1 expressions were significantly negatively correlated with FCR (r = -0.81 to -0.93; P < 0.01), whereas PPARγ and FAS expressions were significantly positively correlated with FCR (r = 0.74 to 0.87; P < 0.01). PPARγ expression was significantly positively correlated with RFI (r = 0.83; P < 0.01), whereas CPT1A and ACOX1 expressions were significantly negatively correlated with RFI (r = -0.84 to -0.89; P < 0.01). Sirt1 mRNA expression was positively correlated with Foxo1, CPT1A, and ACOX1 mRNA expression (r = 0.78 to 0.92; P < 0.01). Association of Foxo1 with CPT1A and ACOX1 was positive (r = 0.88 to 0.96; P < 0.01). These results suggest that genes related to fatty acid oxidation are upregulated in the liver of ducks with high feed efficiency, while genes associated with lipid synthesis are downregulated. Furthermore, the inclusion of lipid metabolism-related genes in future breeding programs might be beneficial for selecting ducks with greater feed efficiency phenotype.

AdipoR1 and AdipoR2 maintain membrane fluidity in most human cell types and independently of adiponectin

The FA composition of phospholipids must be tightly regulated to maintain optimal cell membrane properties and compensate for a highly variable supply of dietary FAs. Previous studies have shown that AdipoR2 and its homologue PAQR-2 are important regulators of phospholipid FA composition in HEK293 cells and Caenorhabditis elegans, respectively. Here we show that both AdipoR1 and AdipoR2 are essential for sustaining desaturase expression and high levels of unsaturated FAs in membrane phospholipids of many human cell types, including primary human umbilical vein endothelial cells, and for preventing membrane rigidification in cells challenged with exogenous palmitate, a saturated FA. Three independent methods confirm the role of the AdipoRs as regulators of membrane composition and fluidity: fluorescence recovery after photobleaching, measurements of Laurdan dye generalized polarization, and mass spectrometry to determine the FA composition of phospholipids. Furthermore, we show that the AdipoRs can prevent lipotoxicity in the complete absence of adiponectin, their putative ligand. We propose that the primary cellular function of AdipoR1 and AdipoR2 is to maintain membrane fluidity in most human cell types and that adiponectin is not required for this function.

Highlights of This Issue

Chen and Zhang and colleagues describe a systematic analysis of molecular correlates of metastasis using The Cancer Genome Atlas (TCGA) datasets across 11 different cancer types, with data involving mRNA, microRNA, protein, and DNA methylation. While some differential expression patterns associated with metastasis may be shared across multiple cancer types, each one also shows a metastasis signature that is distinct. When combining TCGA data with that from multiple independent profiling datasets of metastatic tumors, the resulting catalog of gene-level alterations, according to cancer type, provides a resource for future studies.Cytotoxic T lymphocytes (CTL) use the Fas-mediated apoptosis pathway as one of the two effector mechanisms to suppress tumors. Xiao et al. report here that the CD133+CD24loFaslo colon cancer cell subset exhibits decreased sensitivity to FasL-induced apoptosis in vitro and increased resistance to both tumor-specific CTL adoptive transfer and PD-1 blockade antibody immunotherapies in preclinical mouse tumor models in vivo. Furthermore, Fas expression level is positively correlated with colon cancer patient survival time. This study indicates that the CD133+CD24loFaslo colon cancer cells may, at least in part, underlie colon cancer resistance to immune checkpoint inhibitor immunotherapy.Protocadherins (PCDHs) are transmembrane proteins and members of the Cadherin superfamily that play well-established roles in cell adhesion and regulation of signaling pathways. Both oncogenic and tumor suppressive roles have been assigned to PCDHs. However, the roles of individual PCDHs in cancer and the mechanisms through which their gain-of-function and loss-of-function drive tumorigenesis in vivo remain poorly understood. The current study establishes a critical oncogenic function for PCDH7 in lung cancer using a novel transgenic mouse model. Moreover, somatic genome editing of Pcdh7 with CRISPR/Cas9 in KrasLSL-G12D; Tp53fl/fl mice support the potential therapeutic efficacy of PCDH7 inhibition for patients with lung cancer.Prostate cancer can develop resistance to anti-androgen therapy by reactiving Androgen Receptor (AR) signaling. The critical processes that lie downstream of AR and contribute to tumor growth and therapeutic resistance remain to be fully defined. Zennami and colleagues identified PDCD4, a tumor suppressive protein, as a key target of AR signaling in prostate cancer cells. AR signaling suppresses PDCD4 expression, through miR-21, leading to enhanced cell proliferation, survival, tumorigenesis, and castration resistance. These results uncover PDCD4 loss as a new molecular mechanism of castration resistance and suggest that pathways downstream of PDCD4 may be novel targets for therapy.

Expression of the apoptosis-releated genes in patients with newly diagnosed chronic lymphocytic leukemia in clinical data context

Introduction.The given data of fundamental studies of apoptosis processes in B-cell lymphocytic leukemia (B-CLL) testifies about the complexity and variety of mechanisms affecting the kinetics of normal cells and tumor lymphocytes in this disease. It is important to study the severity of clinical manifestations of the disease depending on the expression of the genes that modulate apoptosis.The purposeof the study is to compare the activity of genes encoding apoptosis modulators, the cell cycle and cancer-testicular PRAME protein with clinical manifestations of the disease in primary patients with B-CLL.Materials and methods.The level of expression of the proapoptotic genes FAS, TRAIL, TNFR2, DR4/5 and DR3, as well as the HSP27, XIAP genes, blocking apoptosis was determined in 23 patients with newly diagnosed chronic B-CLL. In addition, expression of genes TP53 and P21 and cancer-testis gene PRAME are tested.Results.According to the multivariate regression analysis, the FAS gene expression in the onset of the disease had the greatest impact on the clinical characteristics of the disease. In this connection, the patients were divided into groups with normal (group) and low gene level (group II). A low level of FAS expression (Me 387 %) was associated with stage II disease (p = 0.03), a large number of lympho cytes (p = 0.001), fewer erythrocytes (p = 0.08), and a lower level of TNFR2 gene expression (p = 0.08), high level of expression of XIAP, HSP27, P21. Overall, the anti-apoptotic potential in Group II patients was higher, which was accompanied by more pronounced clinical manifestations of the disease.Conclusions.The increased anti-apoptotic potential of tumor lymphocytes in newly diagnosed B-CLL is accompanied by a larger tumor mass and greater clinical and hematological manifestation of the disease.

Effects of dietary n-3 LC-PUFA/n-6 C18 PUFA ratio on growth, feed utilization, fatty acid composition and lipid metabolism related gene expression in black seabream, Acanthopagrus schlegelii

An 8-week feeding trial with black seabream, Acanthopagrus schlegelii, was conducted to evaluate effects of dietary n-3 LC-PUFA/n-6 C18 PUFA (LC-PUFA, long-chain polyunsaturated fatty acid; C18 PUFA, PUFA with 18 atoms of carbon) ratio on growth, feed utilization, fatty acid composition and lipid related gene expression. Five isoproteic and isolipidic diets were formulated to contain different n-3 LC-PUFA/n-6 C18 PUFA ratios: 0.1, 0.4, 0.5, 0.8, 1.0. Each diet was assigned randomly to feed triplicate net cage of 90 fish (30 fish per net cage) of initial weight 3.0 ± 0.0 g. As a result, final weight (FW), weight gain (WG), specific growth rate (SGR), feed efficiency (FE) and protein efficiency ratio (PER) were all significantly affected by dietary n-3 LC-PUFA/n-6 C18 PUFA ratio (P < 0.05). Fish fed the diet with a n-3 LC-PUFA/n-6 C18 PUFA ratio of 0.8 showed the highest FW, WG, SGR, FE, PER. There was no significant differences in survival rate (SR), hepatosomatic index (HIS), viscerosomatic index (VIS), intraperitoneal fat ratio (IPF) among all treatments (P > 0.05). Likewise,The moisture contents, crude protein, crude lipid and ash contents in the whole body and muscle were all not affected by experimental diets (P > 0.05). Tissue fatty acid profiles reflected the diets. The highest concentration of triacylglycerol (TG) was observed in fish fed the lowest ratio (0.1) (P < 0.05). However,the highest activities of alanine aminotransferase (ALT) and aspartate transaminase (AST) as well as the highest content of cholesterol (CHOL) were all found in fish fed the diet containing n-3 LC-PUFA/n-6 C18 PUFA ratio of 1.0 (P < 0.05). Hepatic G6pd, Lpl and Hsl activities were significantly affected by dietary n-3 LC-PUFA/n-6 C18 PUFA ratio (P < 0.05). Similarly, the expression levels of fas, accɑ, g6pd, cpt1ɑ and atgl were influenced significantly among treatments, the highest fas, accɑ, g6pd, cpt1ɑ and atgl expression levels were obtained in fish fed the lowest dietary n-3 LC-PUFA/n-6 C18 PUFA ratio of 0.1 (P < 0.05). In conclusion, according to quadratic regression analysis, the optimum dietary n-3 LC-PUFA/n-6 C18 PUFA ratio was 0.7 for juvenile black seabream to obtain the maximum growth. Moreover, this study also indicated that dietary n-3 LC-PUFA/n-6 C18 PUFA ratio affected fatty acid profiles and significantly influenced lipid metabolism.

Characteristics of peripheral V\u03b32V\u03b42 T cells in interferon-\u03b3 release assay negative pulmonary tuberculosis patients

BackgroundIt is not fully explained why some active tuberculosis patients show negative interferon-γ release assays (IGRAs). In this study, we tried to explore associations of IGRAs with the characteristics of peripheral Vγ2Vδ2 T cells and their functions of producing cytokines.Methods32 pulmonary tuberculosis patients were enrolled and divided into two groups according to their IGRAs results: 16 with IGRA-negative as test group and 16 with IGRA-positive as control group. Chest X-rays and T-SPOT.TB tests were performed and the severity of the lung lesions was scored. The amount of Vγ2Vδ2T cell and their expression levels of the apoptosis-related membrane surface molecule Fas and FasL in peripheral blood were analyzed by flow cytometry, and the function of secreting cytokines (IFN-γ, TNF-α and IL-17A) of Vγ2Vδ2 T cell were determined by intracellular cytokine staining.ResultsThe IGRA-negative TB patients had more lesion severity scores and displayed reduced peripheral blood Vγ2Vδ2 T cell counts (p = 0.009) as well as higher Fas and FasL expression in peripheral blood Vγ2Vδ2 T cells (p = 0.043, 0.026). A high lesion severity score was correlated with a decreased Vδ2+ T cell number and increased Vγ2Vδ2 T cells Fas/FasL expression leve in the peripheral blood (p = 0.00, P < 0.01). The function of secreting cytokines was slightly impaired in IGRA-negative TB patients (p = 0.402). There is no significant differences in expression levels of Fas and FasL in CD4+ T cells (p = 0.224, 0.287) or CD8+ T cells (p = 0.184, 0.067) between test and control groups.ConclusionCompared with IGRA-positive TB patients, the IGRA-negative TB patients had more lesion severity scores, the number of Vγ2Vδ2 T cells decreased and the function of secreting cytokines impaired. In addition, we suggest that increased expression of Fas/FasL triggers Vγ2Vδ2 T cell apoptosis.

Propranolol induces hemangioma endothelial cell apoptosis via a p53‑BAX mediated pathway.

The use of propranolol for the treatment of infantile hemangioma (IH) has been widely investigated in recent years. However, the underlying therapeutic mechanism of propranolol for the treatment of IH remains poorly understood. The aim of the present study was to investigate the expression of proteins regulated by cellular tumor antigen p53 (p53) in associated apoptosis pathways in IH endothelial cells (HemECs) treated with propranolol. Furthermore, the present study aimed to investigate the exact apoptotic pathway underlying the therapeutic effect of propranolol against IH. In the present study, HemECs were subcultured and investigated using an inverted phase contrast microscope, immunocytochemical staining and a scanning electron microscope (SEM). Experimental groups and blank control groups were prepared. All groups were subjected to drug treatment. A high p53 expression model of HemECs was successfully established via transfection, and a low p53 expression model of HemECs was established using pifithrin-α. The apoptosis rate of each group was determined using Annexin V-fluorescein isothiocyanate/propidium iodide double staining and flow cytometry. The expression levels of downstream proteins regulated by p53 [tumour necrosis factor receptor superfamily member 6 (FAS), p53-induced death domain-containing protein (PIDD), death receptor 5 (DR5), BH3-interacting domain death agonist (BID), apoptosis regulator BAX (BAX), p53 unregulated modulator of apoptosis (PUMA), phosphatidylinositol-glycan biosynthesis class S protein (PIGS), and insulin-like growth factor-binding protein 3 (IGF-BP3)] were revealed in the experimental and control groups via western blotting. Microscopic observation revealed the growth of an adherent monolayer of cells, which were closely packed and exhibited contact inhibition. Immunocytochemical staining demonstrated increased expression of clotting factor VIII. SEM analysis revealed presence of Weibel-Palade bodies. The results of the analyses verified that the cultured cells were HemECs. The staining of the samples resulted in a significantly increased rate of apoptosis in experimental groups compared with the blank control group. This result suggested that there is an association between p53 expression and the rate of apoptosis of propranolol-treated HemECs. The results of the western blot analysis demonstrated an upregulation of BAX expression and a downregulation of IGF-BP3 expression in the HemECs treated with propranolol. There were no significant differences in the expression levels of FAS, DR5, PIDD, BID, PUMA and PIGS between experimental and control groups. This result suggests that p53 has an important role in HemEC apoptosis. The results of the present study additionally suggest that the propranolol-induced HemEC apoptosis pathway is a mitochondrial apoptosis pathway and is regulated by p53-BAX signaling.