Oleic acid-induced steatosis model establishment in LMH cells and its effect on lipid metabolism.

Hepatic steatosis is a highly prevalent liver disease, yet research on it is hampered by the lack of tractable cellular models in poultry. To examine the possibility of using organoids to model steatosis and detect it efficiently in leghorn male hepatocellular (LMH) cells, we first established steatosis using different concentrations of oleic acid (OA) (0.05-0.75 mmol/L) for 12 or 24 h. The subsequent detections found that the treatment of LMH cells with OA resulted in a dramatic increase in intracellular triglyceride (TG) concentrations, which was positively associated with the concentration of the inducing OA (R2 > 0.9). Then, the modeled steatosis was detected by flow cytometry after NileRed staining and it was found that the intensity of NileRed-A was positively correlated with the TG concentration (R2 > 0.93), which demonstrates that the flow cytometry is suitable for the detection of steatosis in LMH cells. According to the detection results of the different steatosis models, we confirmed that the optimal induction condition for the establishment of the steatosis model in LMH cells is OA (0.375 mmol/L) incubation for 12 h. Finally, the transcription and protein content of fat metabolism-related genes in steatosis model cells were detected. It was found that OA-induced steatosis could significantly decrease the expression of nuclear receptor PPAR-γ and the synthesis of fatty acids (SREBP-1C, ACC1, FASN), increasing the oxidative decomposition of triglycerides (CPT1A) and the assembly of low-density lipoproteins (MTTP, ApoB). Sterol metabolism in model cells was also significantly enhanced (HMGR, ABCA1, L-BABP). This study established, detected, and analyzed an OA-induced steatosis model in LMH cells, which provides a stable model and detection method for the study of poultry steatosis-related diseases.

Rothia nasimurium as a Cause of Disease: First Isolation from Farmed Chickens.

Rothia nasimurium is a facultative anaerobic Gram-positive coccus belonging to the Rothia genus of the Micrococcaceae family. While Rothia nasimurium is considered an opportunistic pathogen, to date few studies have investigated its pathogenicity and drug resistance. In January 2022, chickens at a poultry farm in China's Xinjiang Uygur Autonomous Region became ill and died. Treatment with commonly used Chinese medicines and antibiotics was ineffective, causing economic losses to the poultry farm. In order to determine the cause of the disease in these poultry farm chickens, the isolation and identification of the pathogens in the livers and other internal organs of the sick and dead chickens were performed. Further, animal pathogenicity tests, antibiotic susceptibility tests, and the detection of antibiotic resistance genes were carried out to analyze the pathogenicity and drug resistance of the identified pathogens. A Gram-positive coccus was isolated from the livers of the diseased chickens. The isolate was resistant to 17 antibiotics, including ciprofloxacin, chloramphenicol, and florfenicol, and was only sensitive to penicillin, amikacin, and tigecycline, to varying degrees. The results of the drug resistance gene testing indicated that the isolated bacterium carried 13 kinds of resistance genes. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, morphological observations, biochemical tests, and 16S rRNA gene sequence analysis were performed on the isolated bacterium, and it was determined that the isolated bacterial strain was Rothia nasimurium. The animal pathogenicity tests showed that the isolate caused feather loss and death in chicks; the clinical symptoms and necropsy lesions of the test chicks were consistent with those observed in the farmed chickens. A review of the literature revealed that, to date, there are no reports of infection with Rothia nasimurium in chickens. Thus, in this study, Rothia nasimurium was isolated from chickens for the first time and an investigation of the biological characteristics of the bacterium was carried out in order to provide a reference for the clinical treatment, prevention, and control of Rothia nasimurium infection.

[Effect of acupuncture on expression of transfer growth factor-ß1 in lacrimal gland of rabbits with dry eye].

OBJECTIVE: To observe the effect of acupuncture on expression of transforming growth factor-ß1(TGF-ß1) in lacrimal gland of rabbits with dry eye, so as to explore its underlying mechanism in improving dry eye. METHODS: Healthy male New Zealand rabbits were randomly assigned to 5 groups (n=6 in each group), namely, blank group, model group, western medicine group, acupuncture group and sham acupuncture group. The dye eye rabbit model was estabilished by subcutaneous injection of Scopolamine Hydrobromide solution for 21 days. After modeling, rabbits in the western medicine group were treated with Flumirone eye drops in their eyes 3 times a day, one drop each time. Rabbits of the acupuncture group reveived electroacupuncture(4 Hz/20 Hz, 1 mA) at "Cuanzhu"(BL2) and "Tongziliao"(GB1) for 15 min, and received acupuncture at "Jingming"(BL1), "Taiyang" (EX-HN5) and "Sizhukong"(TE23) for 15 min, once a day. Rabbits of the sham acupuncture group received blunt acupuncture at the surface of the same acupoints once a day. All the treatments were conducted for 14 days. The changes of tear flow, tear film break-up time (BUT) and lacrimal gland morphology in each group were observed. The expression of TGF-ß1 protein and mRNA in lacrimal gland were detected by Western blot and quantitative real-time fluorescence PCR respectively. RESULTS: Following modeling, except for the blank group, the tear flow and BUT in other 4 groups decreased significantly (P<0.01). Compared with their own pretreatment, the tear flow and BUT in western medicine group and acupuncture group increased after the treatment (P<0.05). Compared with the model group, the tear flow and BUT increased in the western medicine group and the acupuncture group(P<0.05). Atrophic lacrimal epithelial cells and the stroma of mucous membrane infiltrated by lymphocytes and plasma cells were found in rabbits of the model group and the sham acupuncture group. By contrast, in the western medicine group and the acupuncture group, the structure of lacrimal epithelial cells was basically normal, and the infiltration of lymphocytes and plasma cells were scattered in the stroma of mucous membrane. In comparison with the blank group, the expression of TGF-ß1 protein and mRNA in lacrimal gland were significantly up-regulated in the model and sham acupuncture groups (P<0.01). Compared with the model group, the expression of TGF-ß1 protein and mRNA were significantly down-regulated in the western medicine and acupuncture groups (P<0.01, P<0.05). CONCLUSION: Acupuncture intervention can increase tear flow and BUT in rabbits with dry eye, which may be related to the regulation of TGF-ß1 expression in lacrimal gland.

Trends in global, regional and national incidence of pneumoconiosis caused by different aetiologies: an analysis from the Global Burden of Disease Study 2017.

OBJECTIVES: Pneumoconiosis remains a major global occupational health hazard and illness. Accurate data on the incidence of pneumoconiosis are critical for health resource planning and development of health policy. METHODS: We collected data for the period between 1990 and 2017 on the annual incident cases and the age-standardised incidence rates (ASIR) of pneumoconiosis aetiology from the Global Burden of Disease Study 2017. We calculated the average annual percentage changes of ASIR by sex, region and aetiology in order to determine the trends of pneumoconiosis. RESULTS: Globally, the number of pneumoconiosis cases increased by a measure of 66.0%, from 36 186 in 1990 to 60 055 in 2017. The overall ASIR decreased by an average of 0.6% per year in the same period. The number of pneumoconiosis cases increased across the five sociodemographic index regions, and there was a decrease in the ASIR from 1990 to 2017. The ASIR of silicosis, coal workers' pneumoconiosis and other pneumoconiosis decreased. In contrast, measures of the ASIR of asbestosis displayed an increasing trend. Patterns of the incidence of pneumoconiosis caused by different aetiologies were found to have been heterogeneous for analyses across regions and among countries. CONCLUSION: Incidence patterns of pneumoconiosis which were caused by different aetiologies varied considerably across regions and countries of the world. The patterns of incidence and temporal trends should facilitate the establishment of more effective and increasingly targeted methods for prevention of pneumoconiosis and reduce associated disease burden.

Antioxidant tert-butylhydroquinone ameliorates arsenic-induced intracellular damages and apoptosis through induction of Nrf2-dependent antioxidant responses as well as stabilization of anti-apoptotic factor Bcl-2 in human keratinocytes.

UNLABELLED: Human skin is a known target site of inorganic arsenic with effects ranging from hyperkeratosis to dermal malignancies. Tert-butylhydroquinone (tBHQ), approved food-grade phenolic antioxidant, is demonstrated to induce remarkable antioxidant activity in a variety of cells and tissues. The present study aimed at the protective effects of tBHQ on arsenic-induced cytotoxicity and apoptosis in human keratinocytes. Our results demonstrated that tBHQ antagonized arsenic-induced decrease of cell viability, generation of reactive oxygen species (ROS) and lipid peroxidation, as well as reduction of antioxidative enzymes superoxide dismutase (SOD) and catalase (CAT) activities. We also found that tBHQ relieved the G2/M phase arrest by arsenic exposure, which was associated with altering the expression of cell cycle regulators cyclin D1 and CDK4. tBHQ treatment further reduced the numbers of arsenic-induced mitochondrial-mediated apoptotic cells, which occurred concomitantly with the effective recovery of mitochondrial membrane potential (ΔΨm) depolarization, the release of cytochrome c releasing from the mitochondrial as well as the survival signal related factor caspase 3 activation. Our experiments then confirmed that tBHQ activated nuclear factor E2-related factor 2 (NRF2) pathway by increasing NRF2 protein in both nucleus and cytoplasm and upregulating NRF2 downstream targets NAD(P)H: quinine oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1). More interestingly, arsenic-induced decrease of anti-apoptotic factor B-cell lymphoma-2 (Bcl-2) and increase of pro-apoptotic factor Bcl-2-associated X protein (Bax) could all be reversed by tBHQ pretreatment. These results suggested together that tBHQ could ameliorate arsenic-induced cytotoxicity and apoptosis, which might be linked with the induction of Nrf2-dependent antioxidant responses as well as stabilization of anti-apoptotic factor Bcl-2 in human keratinocytes.

Tert-butylhydroquinone as a phenolic activator of Nrf2 antagonizes arsenic-induced oxidative cytotoxicity but promotes arsenic methylation and detoxication in human hepatocyte cell line.

Oxidative stress plays crucial roles in exerting a variety of damages upon arsenic exposure. Nuclear factor (erythroid-derived 2)-like 2 (Nrf2) is a master transcriptional regulator protecting cells and tissues from oxidative injuries. The objective of this study was to test whether tert-butylhydroquinone (tBHQ), a well-known synthetic Nrf2 inducer, could protect human hepatocytes against arsenic-induced cytotoxicity and oxidative injuries. Our results showed that 5 and 25 µmol/l tBHQ pretreatment suppressed the arsenic-induced hepatocellular cytotoxicity, reactive oxygen species generation, and hepatic lipid peroxidation, while relieved the arsenic-induced disturbances of intracellular glutathione balance. In addition, we also observed that tBHQ treatment promoted the arsenic biomethylation process and upregulated Nrf2-regulated downstream heme oxygenase-1 and NADPH: quinine oxidoreductase 1 mRNA expressions. Collectively, we suspected that Nrf2 signaling pathway may be involved in the protective effects of tBHQ against arsenic invasion in hepatocytes. These data suggest that phenolic Nrf2 inducers, such as tBHQ, represent novel therapeutic or dietary candidates for the population at high risk of arsenic poisoning.

[Effects of inorganic arsenic on mRNA expression of Nrf2 and Nrf2-regulated downstream anti-oxidant enzymes in Chang hepatocytes].

OBJECTIVE: To study the effects of inorganic arsenic on mRNA expression of transcription factor Nrf2 and Nrf2-regulated downstream anti-oxidant enzymes, NADPH: quinine oxidoreductase 1 (NQO1) and heme oxygenase 1(HO-1) in Chang hepatocytes. METHODS: Chang hepatocytes were treated with 5, 10, 25 and 50 micromol/L of sodium arsenite (NaAsO2) for 6h, and RT-PCR were then performed to detect the mRNA expression of Nrf2, NQO1 and HO-1. RESULTS: When exposed to 5, 10, 25 and 50 micromol/L of NaAsO2, Nrf2 mRNA were (100.74 +/- 3.70)%, (105.96 +/- 1.75)%, (101.76 +/- 1.01)% and (101.81 +/- 6.33)% of control,showing no statistic significance (P > 0.05). NQO1 mRNA expression were significantly increased (P < 0.05), and the levels of NQO1 mRNA were (106.52 +/- 3.11)%, (113.27 +/- 2.84)%, (111.96 +/- 6.96)% and (107.33 +/- 2.76)% of control, respectively. Moreover, HO-1 mRNA expression were also significantly induced by sodium arsenite exposure, and a definite dose-effect relationship was confirmed (P < 0.01). CONCLUSION: No elevation of Nrf2 transcription was found, while inorganic arsenic could increase the Nrf2-regulated NQO1 and HO-1 mRNA expression in human hepatocytes.