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ObjectiveTo explore the cytotoxicity of four wild mushrooms involved in a case of Yunnan sudden unexplained death （YNSUD）, to provide the experimental basis for prevention and treatment of YNSUD.MethodsFour kinds of wild mushrooms that were eaten by family members in this YNSUD incident were collected and identified by expert identification and gene sequencing. Raw extracts from four wild mushrooms were extracted by ultrasonic extraction to intervene HEK293 cells, and the mushrooms with obvious cytotoxicity were screened by Cell Counting Kit-8 （CCK-8）. The selected wild mushrooms were prepared into three kinds of extracts, which were raw, boiled, and boiled followed by enzymolysis. HEK293 cells were intervened with these three extracts at different concentrations. The cytotoxicity was detected by CCK-8 combined with lactate dehydrogenase （LDH） Assay Kit, and the morphological changes of HEK293 cells were observed under an inverted phase contrast microscope.ResultsSpecies identification indicated that the four wild mushrooms were Butyriboletus roseoflavus, Boletus edulis, Russula virescens and Amanita manginiana. Cytotoxicity was found only in Amanita manginiana. The raw extracts showed cytotoxicity at the mass concentration of 0.1 mg/mL, while the boiled extracts and the boiled followed by enzymolysis extracts showed obvious cytotoxicity at the mass concentration of 0.4 mg/mL and 0.7 mg/mL, respectively. In addition to the obvious decrease in the number of HEK293 cells, the number of synapses increased and the refraction of HEK293 cells was poor after the intervention of Amanita manginiana extracts.ConclusionThe extracts of Amanita manginiana involved in this YNSUD case has obvious cytotoxicity, and some of its toxicity can be reduced by boiled and enzymolysis, but cannot be completely detoxicated. Therefore, the consumption of Amanita manginiana is potentially dangerous, and it may be one of the causes of the YNSUD.

Computed tomography (CT) examination is the major measure for detecting and diagnosis of foreign bodies in human body. Although CT has high sensitivity in diagnosis of foreign body, some interference factors may still lead to missed diagnosis or misdiagnosis. Here we report a rare case that a bamboo stick accidentally pierced into the left chest of a young man who was drunk and unware of this hurt. The patient experienced cough, chest pain, fever, hemoptysis, and was misdiagnosed as primary and secondary tuberculosis based on chest CT examinations at a local hospital, although no tubercular bacillus detected by sputum smear. He subsequently received anti-tuberculous treatments in the following three years, but no improvement of his symptoms was observed. Until one month before his death, the bamboo stick was detected by spiral CT examination as well as three-dimensional image reconstruction at another hospital. Postmortem examination revealed pneumonia, pulmonary infarction, and abscess as the causes of his death. We analyze the potential reasons of misdiagnosis in this case, aiming to provide reference for the diagnosis and treatment of pulmonary inflammation associated with foreign body in the future.

Aim: To study the expressions of p27 kip1 protein and p27mRNA, the hypermethylation of p27 kip1 and the relation between them in various stages of hepatocarcinogenesis., Methods: p27 protein and p27mRNA were detected by immunohistochemical staining and in situ hybridization respectively in 68 cases of normal liver, liver cirrhosis, pericancerous cirrhosis and hepatocellular carcinoma (HCC). The hypermethylation of p27 kip1 was detected by methylation-specific PCR (MSP) in 44 cases of normal liver, liver cirrhosis, and HCC., Results: The positive rate of p27 protein was 66.7% (4/6) in normal liver, 60.0% (6/10) in liver cirrhosis, 50.0% (12/24) in pericancerous cirrhosis and 21.4% (6/28) in HCC. There were no statistical differences in normal liver, liver cirrhosis and pericancerous cirrhosis, but the positive rate of p27 protein significantly decreased in HCC compared to that in the other groups (P = 0.006, chi2 = 7.664). The positive rate of p27 kip1 mRNA was 83.3% (5/6) in normal liver, 70.0% (7/10) in liver cirrhosis, 75.0% (18/24) in pericancerous cirrhosis and 25.0% (7/28) in HCC. There were no statistical differences in normal liver, liver cirrhosis and pericancerous cirrhosis, but the positive rate of p27 kip1 mRNA also significantly decreased in HCC compared to that in the other groups (P = 0.000, chi2 = 16.600). In addition, there was a significant correlation between the expression of p27 protein and p27mRNA in the integrated group of normal liver and liver cirrhosis. However, no significant correlation was found between pericancerous cirrhosis and HCC. Using MSP, we found that 1 HCC in 44 cases (including 6 cases of normal liver, 10 cases of liver cirrhosis and 28 cases of HCC) was methylated, whose p27 protein and p27mRNA were negative., Conclusion: The reduction or loss of p27 protein and p27mRNA are potentially involved in hepatocarcinogenesis. The hypermethylation of p27 might lead to the loss of p27mRNA transcription.