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Objective:To investigate the efficacy of metagenomic next generation sequencing (mNGS) in the etiological diagnosis of patients with spinal infection, so as to provide reference for timely diagnosis and treatment.Methods:A total of 40 patients with suspected spinal infection admitted to the Department of Infectious Diseases in Henan Provincial People′s Hospital from January 2020 to July 2022 were included. The results of tissue culture, histopathological examination and tissue mNGS detection were analyzed retrospectively. According to the clinical diagnose, the patients were divided into the spinal infection group (28 cases) and the non-spinal infection group (12 cases). The positive rate, sensitivity and specificity of mNGS and tissue culture in the pathogen detection of patients with spinal infection were compared. McNemar test was used for statistical analysis.Results:There were 23 males and 17 females in 40 patients. The positive rate of mNGS was higher than that of tissue culture (75.0%(30/40) vs 12.5%(5/40)), and the difference was statistically significant ( χ2=0.08, P<0.001). Based on clinical diagnostic criteria, the sensitivity of mNGS in the diagnosis of spinal infection was higher than that of tissue culture (82.1% vs 17.9%), with a statistically significant difference ( χ2=0.02, P<0.001), while the specificity compared to the tissue culture (33.3% vs 100.0%), the difference was not statistically significant ( P>0.05). Conclusions:mNGS has a high pathogen detection rate and sensitivity in the etiological diagnosis of patients with spinal infection, which could provide clinical guidance for the diagnosis and treatment of patients with spinal infection.
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Objective To investigate the current status of hepatitis B virus (HBV) infection among hospitalized patients aged 1-18 years, as well as the status of immunity after hepatitis B vaccination. Methods Related data were collected from the patients aged 1-18 years who were hospitalized in Henan Provincial People's Hospital from July 2020 to July 2021, including serological markers for hepatitis B (HBsAg, anti-HBs, HBeAg, anti-HBe, and anti-HBc) and hepatitis B vaccination. The epidemiological situation of HBV infection was analyzed, as well as the immune effect after vaccination. The trend chi-square test was used for trend analysis. Results A total of 10 658 hospitalized patients were collected, among whom there were 6 372 male patients (59.79%) and 4 286 female patients (40.21%). In this population, the patients with positive HBsAg accounted for 0.28% (30/10 658), with a relatively high proportion of 0.68% and 0.62%, respectively, in the 17-and 18-year age groups; the patients with positive anti-HBs accounted for 51.82% (5 523/10 658), with a relatively high proportion of > 63% in the 1-4 years age groups, and there was a reduction in the proportion of patients with positive anti-HBs (fluctuating around 40%) in the 5-18 years age groups. With the increase in age, the positive rate of anti-HBs tended to decrease in both male and female patients (male: χ 2 =8.217, P =0.004; female: χ 2 =10.048, P =0.002). Conclusion Based on the data of hospitalized patients, HBV infection in the population aged 1-18 years in Henan Province has the characteristics of low prevalence rate and high immunity, and the reduction in the proportion of patients with positive anti-HBs at more than five years after vaccination should be taken seriously in this region.
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In recent years, the continuous updating of medical means and the rapid development of assisted reproductive technology provide an effective way for cancer patients to preserve and restore fertility, but it still face many challenges in the specific application process. In view of the status quo of fertility preservation of cancer patients at home and abroad, factors affecting fertility of cancer patients such as radiotherapy, chemotherapy and surgery, and common fertility preservation methods of cancer patients such as embryos, oocytes, sperm cryopreservation, ovarian transplantation and other technologies, this paper considered related ethical issues from the following aspects: following the informed consent of cancer patients, executing the relevant laws and regulations on fertility preservation of cancer patients, and implementing the whole process supervision of the ethics committee, so as to provide ideas for the follow-up research on fertility preservation of cancer patients.
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Objective@#To observe ascitic interleukin-7 expression level in cirrhotic patients complicated with spontaneous bacterial peritonitis, and to detect the effect of recombinant human IL-7 on CD4+ and CD8+T lymphocyte function.@*Methods@#A total of 84 patients with liver cirrhosis who were hospitalized from August 2017 to April 2018 were selected. Among them, 51 cases were complicated with cirrhosis and untainted ascites, and 33 cases were cirrhosis complicated with spontaneous bacterial peritonitis. Peripheral blood and ascites were collected routinely. The levels of IL-7 in peripheral blood and ascites were measured by enzyme-linked immunosorbent assay. CD4+T cells and CD8+T cells were purified from ascites, and were stimulated with recombinant IL-7. Cellular proliferation, key transcription factors for mRNA, and cytokines production by CD4+T cells in response to IL-7 stimulation was measured. mRNA expression corresponding to perforin, granzyme B, and granulysin as well as cytokines production by CD8+T cells was also measured in response to IL-7 stimulation. Cytolytic and non-cytolytic activity of CD8+T cells in response to IL-7 stimulation was also investigated in both direct and indirect contact co-culture system. Measurement data of the normal distribution were compared between the two groups by Student’s t-test and the data before and after stimulation were compared by paired t-test. Measurements that did not conform to normal distribution were compared between the two groups using Mann-Whitney U test, and data before and after stimulation were compared using Wilcoxon paired test.@*Results@#There was no significant statistical difference in serum IL-7 levels between the two groups [(5 001 ± 1 458) pg/ml vs. (4 768 ± 1 128) pg/ml, P = 0.41]. The level of ascitic IL-7 in cirrhotic patients complicated with SBP was significantly lower than cirrhosis patients with untainted ascites [(966.4 + 155.8) pg/ml vs. (792.1 + 126.4) pg/ml, P < 0.01]. Recombinant IL-7 stimulation promoted the proliferation of CD4+ and CD8+T cells from ascites in patients with liver cirrhosis complicated by SBP. T-bet mRNA relative expression and IFN-γ secretion in CD4+T cells was also elevated in response to IL-7 stimulation in vitro. Moreover, IL-7 stimulation also increased the mRNA expressions of perforin, granzyme B, and granulysin as well as productions of IFN-γ and TNF-α by CD8+T cells. Recombinant IL-7 stimulation elevated cytolytic and non-cytolytic activity of CD8+T cells from ascites in patients with liver cirrohosis complicated by SBP, which manifested as increased target cell death and IFN-γ production in both direct and indirect contact co-culture system.@*Conclusion@#Ascitic IL-7 promotes T lymphocyte function in patients with liver cirrhosis complicated with SBP.
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Objective To explore the molecular mechanisms of hepatic stimulator substance (HSS) gene knockout in promoting the development and progression of nonalcoholic steatohepatitis (NASH).Methods NASH model mice (n=20) with HSS wild-type (HSS+/+) or HSS gene knockout (HSS-/-) were constructed using modified choline-deficient diet (CD-diet),untreated C57BL6-HSS-/-and C57BL6-HSS+/+ mice (n=20) were considered as control.Ten mice of each group were killed at month 1 and 2,respectively.The levels of triglyceride (TG) and total cholesterol (TC) in liver were measured using ELISA method.Histopathology and collagen deposition in liver tissue were observed using HE staining and Masson staining,respectively.Lipid content in liver tissue was observed and calculated using oil red O staining.The levels of mRNA and proteins of peroxisome proliferators activated receptor gama coactivator 1 alpha (PGC-1α),mitochondrial transcription factor A (TFAM),transcription factor-E2 related factor α (Nrf2),[-loop,dynamin-related protein 1 (Drp1),mitochondrial fission 1 protein (Fis1),mitofusins 1 (Mfn1),autophagy related gene 3 (Atg3) in liver tissue were detected using Real-time PCR and Western blot,respectively.Content of malonaldehyde (MDA),cyclooxygenase Ⅳ (COX Ⅳ) and adenosine tirphosphate (ATP) were measured using kits,and the activity of respiratory chain complex Ⅴ and cytochrome C oxidase in liver tissue were measured using spectrophotometry.the comparison between groups was done by t test.Results The levels of HSS mRNA and protein in mice-HSS-/-were 0.154± 0.04 and 0.08± 0.01,respectively,which were both significantly lower than those in mice-HSS+/+ (0.952 ± 0.08 and 1.362±-0.130,respectively),and t he differences had statistical significance (t =10.244 and 10.375,respectively,both P＜0.05).One month and 2 months after NASH modeled,TC contents in mice-HSS-/ were (248.6±21.5) μmol/g and (217.4±18.0) μmol/g,respectively,which were both remarkably higher than those in mice-HSS+/+ [(153.5 ± 11.2) μmol/g and (140.8 ±7.5) μmol/g,respectively],and the differences had statistical significance (t=15.270 and 10.524,respectively,both P＜0.05).The results form HE staining,oil red O staining and Masson staining indicated that fat deposition,collage deposition and inflammation in liver tissues of mice-HSS-/-were severer than those in mice-HSS+/+.One month after NASH modeled,protein levels of Drp1,Fis1,Mfn1 and Atg3 in liver tissues of mice-HSS-/ were all significantly decreased compared with those in mice-HSS+/+,and the differences had statistical significance (t=10.705,24.072,9.892 and 17.540,respectively,all P＜ 0.05).Two months after NASH modeled,protein levels of Drp1,Fis1,Mfn1and Atg3 in liver tissues of mice-HSS-/ were all significantly decreased compared with those in mice-HSS+/+,and the differences had statistical significance (t=125.378,15.926,34.330 and 13.437,respectively,all P＜0.05).One month after NASH modeled,mRNA levels of Drp1,Fis1,Mfn1 and Atg3 in liver tissues of mice-HSS-/-were all significantly decreased compared with those in mice-HSS+/+,the differences had statistical significance (t=36.337,40.825,33.508 and 28.104,respectively,all P＜0.05).Two months after NASH modeled,mRNA levels of Drp1,Fis1,Mfn1 and Atg3 in liver tissues of mice-HSS-/-were all significantly decreased compared with those in mice-HSS+/+,and the differences had statistical significance (t=35.210,42.375,27.753 and 20.560,respectively,all P＜0.05).The protein levels of PGC-1α,TFAM,Nrf2 and D-loop in liver of C57BL6-HSS-/-group were lower than those in liver of C57BL6-HSS+/+ group,and the differences had statistical significance (one month:t=20.548,31.036,19.445 and 10.974,respectively;two months:t=9.887,13.330,22.375 and 18.903,respectively,all P＜0.05).The mRNA levels of PGC-1α,TFAM,Nrf2 and D-loop in liver of C57BL6-HSS-/-group were all lower than those in C57BL6-HSS+/+ group,and the differences had statistical significance (one month:t=9.087,12.582,21.451 and 7.774,respectively;two months:t=23.758,17.924,9.924 and 15.209,respectively,all P＜0.05).One month and 2 months after NASH modeled,the levels of ATP mRNA in liver of C57BL6-HSS / group were both significantly lower than those in C57BL6-HSS+/+,and the differences had statistical significance 0=43.775 and 28.375,respectively,both P＜0.05);the levels of COXⅣ mRNA in liver of C57BL6-HSS / group were 0.142 ± 0.06 and 0.068± 0.001,respectively,which were both significantly lower than those in C57BL6-HSS+/+ group (0.255± 0.08 and 0.172 ±0.06,respectively),and the differences had statistical significance (t=28.337 and 19.782,respectively,both P＜0.05);the levels of MDA mRNA in liver of C57BL6-HSS-/-group were 0.973 ±0.112 and 1.253±0.054,respectively,which were both significantly lower than those in C57BL6-HSS+/+ group (0.366±0.02 and 0.872±0.05,respectively),and the differences had statistical significance (t=8.357 and 6.582,respectively,both P＜0.05).Conclusion Deletion of HSS accelerates NASH progression via inhibiting mitochondrial fusion,which leads to dysfunction of mitochondrial respiratory chain and inhibition of fatty acid oxidation.
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ObjectiveTo explore the effects of hepatitis B virus X protein (HBx) on the expression and promoter methylation of the p16 tumor suppressor gene, and to investigate the epigenetic role of HBx in the development and progression of hepatitis B virus (HBV)-associated hepatocellular carcinomas (HCC). MethodsExperiments were performed in the human hepatoblastoma cell line HepG2, HepG2 cells expressing green fluorescent protein (HepG2/GFP), and HepG2 cells stably expressing GFP-HBx fusion protein (HepG2/GFP-HBx). Western blot was used to determine the expression levels of the p16 protein in HepG2 cells, HepG2/GFP cells, and HepG2/GFP-HBx cells. HepG2/GFP-HBx cells were treated with a universal inhibitor of DNA methyltransferase (DNMT), 5-aza-2'-deoxycytidine (5-aza-2'-dC). Methylation-specific polymerase chain reaction (MSP) was used to determine the promoter methylation of the p16 tumor suppressor gene in HepG2 cells, HepG2/GFP cells, and HepG2/GFP-HBx cells treated with or without 5-aza-2′-dC. Multiple-group comparison was made by analysis of variance. ResultsAccording to the results of Western blot, HepG2/GFP-HBx cells had a significantly lower expression level of the p16 protein than HepG2 cells and HepG2/GFP cells (P=0.0007; P=00014); there was no significant difference in the expression level of the p16 protein between HepG2/GFP and HepG2 cells (P＞0.05). The MSP assay revealed partial CpG methylation in the p16 promoter region in HepG2/GFP-HBx cells. No promoter methylation was detected in HepG2 cells or HepG2/GFP cells. Non-methylation in the p16 promoter region was restored in HepG2/GFP-HBx cells treated with 5-aza-2′-dC. ConclusionIn the hepatoblastoma cell line, HBx down-regulates the expression of the p16 tumor suppressor gene by inducing methylation in its promoter region. The DNMT inhibitor, 5-aza-2′-dC, restores non-methylation in the p16 promoter region. The reversible modification provides new insights for the treatment and prevention of HBV-associated HCC.