Reduction of natural killer cells is associated with poor outcomes in patients with hepatitis B virus-related acute-on-chronic liver failure.

BACKGROUND AND AIMS: Natural killer (NK) cells are critical innate effectors that respond to viral infections and contribute to immunopathology. Here, we aimed to investigate the role of NK cells in hepatitis B virus-related acute-on-chronic liver failure (HBV-ACLF) and elucidate the underlying mechanism by examining their phenotypic and functional profiles. METHODS: We included patients with HBV-ACLF (n = 37) and chronic hepatitis B (n = 19), and healthy controls (n = 13) in our study. We examined the phenotype and function of different subsets of peripheral NK cells using flow cytometry and RNA-sequencing analysis, and screened liver NK cells using immunohistochemistry. We detected inflammatory cytokines using a Luminex assay. In addition, we analyzed the relationships between these parameters and disease severity. RESULTS: Peripheral NK cells were decreased and characterized by high expression of caspase-3, Ki67, CXCR3, NKG2D, NKp46, CD107a, and GM-CSF, and typified by higher cell migration and immune response by RNA-sequencing analysis in patients with HBV-ACLF than in those with chronic hepatitis B. Accumulations of CXCL-10 and NK cells were found in the liver, and excessive production of CXCL-10 in the peripheral blood contributed to the apoptosis of NK cells in vitro. The decrease in NK cells was associated with the level of HBV DNA and disease severity and had good prognostic performance in predicting the outcome of patients with HBV-ACLF through AUROC analysis. CONCLUSION: NK cells were significantly decreased and showed dysfunction of phenotypic and functional profiles across distinct subsets in the peripheral blood of patients with ACLF. Crosstalk between CXCL-10 and NK cells may mediate the unbalanced distribution of NK cells. Understanding the dysfunction and decrease in NK cells may provide new insights into ACLF pathogenesis.

Simultaneous quantitation of 3ADON and 15ADON chemotypes of DON-producing Fusarium species in Chinese wheat based on duplex droplet digital PCR assay.

Pathogens within Fusarium species are the primary agents of Fusarium head blight (FHB) of wheat, which bring about yield reduction and deoxynivalenol (DON) contamination and are of great concern worldwide. DON-producing Fusarium species can be classified into 3-acetyldeoxynivalenol (3ADON) and 15-acetyldeoxynivalenol (15ADON) chemotypes according to the trichothecene metabolites they produce. The detection of these two chemotypes of pathogens is paramount to the successful implementation of disease management strategies and pathogen-related DON forecasting models. In this study, a duplex droplet digital PCR (duplex ddPCR) assay was developed that allowed for the simultaneous quantitation of 3ADON and 15ADON chemotypes of DON-producing Fusarium species. The assay specificity was tested against 30 isolates of target Fusarium species and several non-target Fusarium species that are frequently isolated from wheat in China. Analyzing 90 wheat samples collected from the North China plain and Yangtze River plain demonstrated that the duplex ddPCR assay coupled with magnetic bead-based DNA extraction was competent for investigating composition of 3ADON and 15ADON chemotypes in Chinese wheat. This assay will be useful for monitoring the epidemic and geographic distribution of 3ADON and 15ADON chemotypes of FHB pathogens, which will help with the disease control and DON management.

Identification and Determination of Rubrofusarin, Rubrofusarin Isomer, and Their Quinone Forms in Grains Using High-Resolution Mass Spectrometry.

Fungi of the genus Fusarium can produce secondary metabolites such as naphthopyrones and naphthoquinones that are toxic and expected to threaten the food and feed safety. In this study, the occurrence of rubrofusarin, rubrofusarin isomer, and their quinone forms in grains was identified and confirmed using ultrahigh-performance liquid chromatography coupled with hybrid quadrupole orbital ion trap mass spectrometry (Q-Orbitrap MS). The quantitation of these compounds in grain samples was also investigated using Q-Orbitrap MS. The results showed the concentrations of rubrofusarin ranged from 3.278 to 33.82 µg/kg, from 0.815 to 61.86 µg/kg, and from 7.362 to 47.24 µg/kg for the maize, rice, and wheat samples, respectively. By comparison, the abundances of their quinone forms were relatively lower, and the concentration of quinone form of rubrofusarin isomer was relatively higher than that of quinone form of rubrofusarin. These compounds were also confirmed to coexist with other known Fusarium mycotoxins. The data-dependent tandem mass spectra obtained from the Q-Orbitrap MS were validated to provide a wealth of valuable information that allowed for advanced data interpretation for solid confirmation of these compounds in grains. To the best of our knowledge, this is the first study that concerns the occurrence and quantitation of rubrofusarin, rubrofusarin isomer, and their quinone forms in grains.

[Effects of long-term fertilization on evolution of S forms in a red soil and a black soil].

Sulfur (S) forms in two contrasting soils (a red soil and a black soil) under different long-term fertilization treatments (from 1990 to 2011) from the National Long-term Monitoring Network of Soil Fertility and Fertilizer Effects of China were investigated using a fractionation scheme in order to explore the distribution and transportation of S with different forms in the soils. The soil samples were collected from the topsoil (0-20 cm) and subsoil (20-40 cm) horizons that were treated with no fertilizers (CK), nitrogen, phosphorus, and potassium fertilizers (NPK), or NPK plus organic manures (MNPK) since 1990. The results indicated that when compared with the CK, total S contents in the topsoil layers treated with NPK and MNPK were increased by 42% and 33% for the red soil, and by 6% and 76% for the black soil, respectively, while the total S in the subsoil layer was less affected by the fertilization treatments and obviously lower than in the topsoil layer except for the red soil treated with NPK. The main forms of inorganic S in the red soil and black soil were found to be available S and HCl-extracted S, respectively. The application of NPK and MNPK increased the available S by 447% and 102% in the topsoil layer of the red soil compared with CK, and facilitated the transportation of available S into the lower depth. In contrast, NPK and MNPK only increased the available S by 54% and 93% in the topsoil layer of the black soil, and showed a slight influence on available S in the subsoil. The organic S forms were predominantly composed of ester S and residual S in the two soils. Under long-term fertilization, the residual S significantly increased over 32% and 55% in the topsoil and subsoil layers, respectively, compared with CK. The ester S and carbon-bonded S, which were relatively active, were less affected by the fertilization treatments, but positively related to the level of organic carbon in each soil (P < 0.05). In addition, the results from the long-term experiments indicated that the contribution of S input from atmospheric deposition was significant and should not be neglected.

[Expressions of CD34 and CD117 in human hepatocellular carcinomas and the clinical significance].

To study the expressions of CD34 and CD117 in the tissues of hepatocelluar carcinoma (HCC) and to explore the relationship with clinical pathology and it's evaluation on the prognosis of HCC patients. The expressions of CD34 and CD117 were examined by two-step methods of PV-9000 of immunohistochemistry in 55 HCC cases, 10 liver cirrhotic specimens and 6 normal liver specimens. Clinical-pathological data, tumor recurrent rate and survival rate after hepatectomy were recorded and analyzed with Fisher's Exact Test, Pearson X2 Test, Kaplan-Meier, Log-Rank Test and Cox Regression. The positive expression of CD34 was found in 65.4% of HCC, 20% of cirrhostic liver specimens and 16.7% of normal liver specimens, respectively. Significant differences found among the three groups, and the CD34 expression was significantly associated with vessel embolus (X2 = 4.000, P = 0.046) and the histological grades (X2 = 11.008, P = 0.001). The positive expression of CD117 was 47.3%, 10% and 0% in HCC, cirrhotic liver specimens and normal liver tissues, respectively, and statistical differences esxisted among the three groups. The CD117 expression was dramatically related to the histological grades (X2 = 5.115, P = 0.024) and clinical stages (X2 = 15.459, P = 0.000). Median disease free survival time after hepatectomy was significantly shorter in the group with positive-expression of CD34 (X2 = 4.105, P = 0.043) and CD117 (X2 = 28.023, P = 0.000) than the negative-expressed groups, respectively. Multivariate analysis showed that CD117 expression status, serum AFP levels and the size of tumor were independently prognostic factors for HCC patients. Tthe results demonstrated that CD34 and CD117 might play an important role in liver carcinogenesis and the progression of HCC, and they might potentially serve as markers for HCC prognosis.

[Transformation of copper and chromium in co-contaminated soil and its influence on bioavailability for pakchoi (Brassica chinensis)].

Heavy metal fractions in soil play an important role in the bioavailability for plants. A pot experiment was carried out to study the transformation and bioavailability for pakchoi of exogenetic copper and hexavalent chromium in co-contaminated soil. The results showed that one month after tested heavy metals were added into soil, the chromium was present mainly in residual and organic bonded form while copper was mainly bound to hydrated oxides of iron and manganese as well as residual form. Lower concentrations copper (< or = 400 mg x kg(-1)) in soil could promote chromium transform from other forms to exchangeable form while higher concentrations copper (> or = 800 mg x kg(-1)) in soil would inhibit this transformation. Similarly, chromium in low concentrations (< or = 5 mg x kg(-1)) could facility copper transform to residual form while chromium in high concentrations (> or = 20 mg x kg(-1)) would reduce such transformation process. Both added chromium to soil polluted by high concentrations copper and added copper to high chromium content soil could promote the polluted soil reach quasi-equilibrium state. Additionally, adding copper in soil could inhibit chromium uptake of pakchoi by promoting soil OM-Cr transformed into RES-Cr. While adding chromium in soil could reduce copper uptake of pakchoi by stimulating the EXE-Cu transformed into the RES-Cu. Therefore, it is necessary to take into consideration the synthetic effects of copper and chromium when carrying on assessment and phytoremediation to co-contaminated soil.

[Correction of the epicanthal fold and angulus oculi using the Z-epicanthoplasty].

OBJECTIVE: To explore a new procedure for aesthetic correction of the medial epicanthal fold aim at the etiopathogenesis. METHODS: The new Z-epicanthoplasty devise the upper and inferior margin of angle of eye medial as one angle of the Z. RESULTS: From 2004 to 2006, 129 patients were treated by using the method. Follow-up 6 to 24 months, all patients were satisfied by eliminating the medial epicanthal fold without obvious scar. CONCLUSIONS: The method is more effect than traditionally Z-plasty. Our technique is a simple, advanced procedure that can be performed widely.

[Correlation of pathology in chronic hepatitis B to viral markers in serum and hepatic tissue].

OBJECTIVE: To investigate the relation of the viral markers in serum and those expressed by hepatocytes to pathological lesions of hepatic tissue in patients with chronic hepatitis B. METHODS: The relation of viral markers including HBsAg, HBsAb, HBeAg, HBeAb, HBcAb and HBV DNA in serum of 647 patients with chronic hepatitis B and HBsAg, HBcAg expressed by hepatocytes in 418 of these patients to pathological lesions of hepatic tissue was determined. RESULTS: Viral markers in serum and those expressed by hepatocytes in patients with chronic hepatitis B were closely correlated with pathological lesions of hepatic tissue. CONCLUSION: The degree of inflammation and fibrosis in hepatic tissue is milder in serum HBsAg, HBeAb, HBcAb positive and HBV DNA negative patients but more serious in those with negative hepatocytic expression of HBsAg and HBcAg. HBV DNA is not significantly associated with pathological lesions of hepatic tissue.

[Reconstruction of the remnant penis: a 52-case report].

OBJECTIVE: To explore the effects of the combined method of abdominal axial flap transposition and penile elongation for the treatment of the remnant penis. METHODS: Fifty-two cases of the remnant penis treated with the combined method from 1984 April to February 2004 were analyzed retrospectively. Follow-up ranged from 0.5 to 20 years postoperatively. RESULTS: The lengths (both in normal and erectile conditions) and the circumferences of the penis gained after operation were (5.6 +/- 1.4) cm, (6.8 +/- 2.5 cm and (6.9 +/- 2.3) cm respectively. The recovery rates of the sensory function were 94.2% and 100% in the glans (immediately and 3 months after operation) and 32.7%, 51.9% and 75% in the flap area (3, 6 and 12 months postoperatively). The two-point distinguishing sense in the glans and the flap area was (5.1 +/- 0.9) mm and(7.9 +/- 1.3) mm 5 years after operation. Early complications included distant flap necrosis (3 cases), disruption of the wound (2 cases), part necrosis of the skin graft in the abdominal wall (2 cases) and poor contours occurred in 4 cases in the later period because of the thickness of the flaps. All of them were corrected with satisfactory results. CONCLUSION: The combined method of abdominal axial flap transposition and penile elongation was recommendable for the treatment of the remnant penis because of its positive effects and less complications.

Pathological changes and clinical manifestations of 1020 children with liver diseases confirmed by biopsy.

BACKGROUND: Liver biopsy plays an important role in accurate diagnosis of various liver diseases in children and liver damages caused by systemic illnesses. This study was designed to evaluate the value of liver biopsy in diagnosis of liver diseases in children and explore the relationship between their pathological changes and clinical manifestations. METHODS: One-second liver biopsy was performed in 1023 pediatric patients with liver diseases at our department from 1983 to 2000. Diagnosis of viral hepatitis was based on the diagnostic criteria formulated by the Chinese Society of Infectious and Parasitic Diseases in 1995. Inflammatory changes of the liver were graded from 0 to 4 (G0-4). RESULTS: Liver biopsy was performed successfully in 1020 patients including 135 infants and young children, of whom 90% were hospitalized patients with chronic liver diseases. Hepatitis virus was the leading cause for chronic liver diseases, among which hepatitis B was detected in 75.4% of the patients. Sixty-nine patients showed liver impairment induced by disorders relevant to that metabolism, Wilson's disease, and glycogen storage disease. Liver inflammatory injury (

[Dynamic expression of matrix metalloproteinase-2, membrane type-matrix metalloproteinase-2 in experimental hepatic fibrosis and its reversal in rat].

OBJECTIVE: To investigate the expression dynamics and significance of matrix metalloproteinase-2 (MMP-2) membrane type-matrix metalloproteinase-2 (MT-MMP-2) in hepatic fibrosis and its reversal counterpart. METHODS: An experimental CCl4 induced hepatic fibrosis rat model was established by intraperitoneal administration of carbon tetrachloride for 2, 4, 6, 8, 10 weeks, and normal rats were used as a control group. The immunohistochemical methods and in situ hybridization were used to detect MMP-2,MT-MMP-2 mRNA and related antigens in the liver. RESULTS: MMP-2,MT-MMP-2 mRNA and related antigens were expressed in mesenchymal cells and parts of hepatocytes besides active pathological changes, especially in the fibrous septum and portal area. Expression of MMP-2,MT-MMP-2 mRNA and related antigens were increased in hepatic fibrosis and decreased gradually in its reversal counterpart. CONCLUSION: This study suggested that mesenchymal cells are the main cellular origins of MMPs. The levels of MMP-2 and MT-MMP-2 antigens and gene expression were closely related to hepatic fibrosis. MMP-2 and MT-MMP-2 may play important roles in hepatic fibrosis and its reversal counterpart.

[Clinical and pathological analysis on characteristics of primary biliary cirrhosis].

OBJECTIVES: To explore the clinical and pathological features and the pathogenesis of primary biliary cirrhosis (PBC) in Chinese Mainland. METHODS: 30 PBC patients were divided into the early group (Scheuer stage I and II, 19 patients) and the late group (Scheuer stage III and IV, 11 patients). The data of clinics and serology were analyzed, and the pathological features of the liver tissues were characterized. The changes of dendritic cells (DCs) and hepatic stellate cells (HSCs) were studied by immunohistochemistry. RESULTS: In all the PBC patients, the rate of the male to the female was 1 to 5, and the average age was 40.6 years. The mean levels of TBiL, ALP and GGT in the sera were (95.9+-88.5) micromol/L, (537.2+-339.2) U/L, and (582.0+-351.2) U/L, respectively. 73.3% patients showed AMA positive, and the level of GGT was positively correlated with the AMA level according to the result of statistical analysis (r=0.778, P=0.000). The symptoms of jaundice and hepatomegaly were presented more commonly in the late group than those in the early group (chi2=5.182, P<0.05; chi2=13.659, P<0.01, respectively). The main changes of morphology of PBC located in portal tracts. The liver tissues in the early stage of PBC showed the damage of bile ducts and obvious proliferation of small bile ducts. The granulomas, the lymphoid follicles and the foamy cells were found in the liver tissues of PBC (2/19 patients, 12/19 patients, and 10/19 patients in the early stage respectively, while 0/11 patients, 4/11 patients, and 3/11 patients in the late stage respectively). There was significant difference between the early stage and the late stage in presence of the lymphoid follicles and the foamy cells (t=4.489, P<0.05; t=4.019, P<0.05, respectively). The biliary pigmentary particles were mainly accumulated in the liver cells around the portal tracts in 90.0% PBC patients, and the accumulation of copper and iron increased, compared with that in normal specimens. The DCs and HSCs located mainly in the portal tracts, especially around the damaged bile ducts. CONCLUSIONS: There are some clinical and pathological characteristics in the patients with PBC. The level of AMA has no direct relationship with the level of transaminase or bilirubin. The proliferated bile ductules may express the antigens which maybe the target of immune attack. As an antigen-presenting cell, DCs may play an important role in the pathogenesis of PBC.

[Clinical pathology and pathogenesis of severe acute respiratory syndrome].

BACKGROUND: To explore the pathological features and pathogenesis of severe acute respiratory syndrome (SARS) to provide evidence for the clinical treatment and prevention of SARS. METHODS: Pathological features of 2 cases of full autopsy and 4 cases of needle biopsy tissue samples from the patients who died from SARS were studied by light and electron microscopy. The distribution and quantity of lymphocyte subpopulations in the lungs and immune organs from SARS patients were analyzed by immunohistochemistry. The location and semi-quantitative analysis of SARS coronavirus in the tissue specimens were studied by electron microscopy, in situ hybridization and immunohistochemistry. RESULTS: In total of 6 cases, diffuse alveolar damage and alveolar cell proliferation were common. The major pathological changes of 2 autopsy cases of SARS in lung tissues were acute pulmonary interstitial and alveolar exudative inflammation, and 2 autopsy and one biopsy lung tissues showed alveolar hyaline membrane formation. Terminal bronchiolar and alveolar desquamation of lung tissues in one autopsy and 2 biopsy cases were noted. Among 6 cases, 2 biopsy cases presented early pulmonary fibrosis and alveolar organization. Meanwhile, the immune organs, including lymph nodes and spleens from 2 autopsy cases of SARS whose disease courses were less than 12 days showed extensive hemorrhagic necrosis, reactive macrophage/histocyte proliferation, with relative depression of mononuclear and granulocytic clones in the bone marrows. However, spleen and bone marrow biopsy tissue samples from 4 dead SARS cases whose clinical course lasted from 21 to 40 days presented repairing changes. SARS coronaviruses were mainly identified in type I and II alveolar epithelia, macrophages, and endothelia; meanwhile, some renal tubular epithelial cells, cardiomyocytes, mucosal and crypt epithelial cells of gastrointestinal tracts, parenchymal cells in adrenal glands, lymphocytes, testicular epithelial cells and Leydig's cells were also detected by electron microscopy combined with in situ hybridization. The semi-quantitative analysis of lymphocyte subpopulations revealed that the proportion of CD8+ T lymphocytes were about 80% of the total infiltrative inflammatory cells in the pulmonary interstitium, with a few CD4+ lymphocytes CD3+, CD4+, CD8+ or CD20+ lymphocyte subpopulations were obviously decreased and there was imbalance in number and proportion, while CD57+, CD68+, S-100+ and HLA-DR+ cells were relatively increased in lymph nodes and spleens. CONCLUSIONS: Histologically, the pulmonary changes could be divided into acute inflammatory exudative, terminal bronchiolar and alveolar desquamative and proliferative repair stages or types during the pathological process of SARS. SARS coronavirus was found in multi-target cells in vivo, which means that SARS coronavirus might cause multi-organ damages which were predominant in lungs. There were varying degrees of decrease and imbalance in number and proportion of lymphocyte subpopulations in the immune organs of the patients with SARS. However, these changes may be reversible. It was found that cellular immune responses were predominant in the lungs of SARS cases, which might play an important role in getting rid of coronaviruses in infected cells and inducing immune mediated injury.

[Cut-off period of subclassification and pathological features of severe hepatitis based on clinical and pathological analyses].

BACKGROUND: To explore the cut-off period of subclassification and pathological features of severe hepatitis (SH). METHODS: Based on combined clinical and pathological analyses, the complete clinical and biopsy or autopsy liver tissues data from 196 cases of patients with severe hepatitis were investigated. Meanwhile, proliferative hepatocytes, cholangioepithelia and collagens were identified by a panel of monoclonal antibodies such as those against albumin, cytokeratin 18,19 and collagen I, III with immunohistochemical method. RESULTS: The clinical and pathological analyses indicated the cut-off periods of acute, subacute and chronic SH (ASH,SSH and CSH) were (13.4+/-7.2) d, (77.4+/-69.3) d and (80.5+/-63.2) d, respectively. Among all SH cases, one case of ASH patient presented clinical manifestation and pathological changes of ASH for 21 days, however, one patient with SSH was demonstrated 12 day course by histological examination. The time of cut-off period between ASH and SSH in child cases was shorter than that in adult cases. Histologically, ASH liver tissues showed massive and/or submassive necrosis caused by one attack, with congestive sinusoid frameworks and proliferative cholangioepithelium-like hepatocytes, while SSH liver tissues presented combined fresh and old submassive or massive necrosis caused by multiple attacks, accompanied by obviously proliferative bile ducts and sinusoid framework collapse.However, the pathological changes of CSH showed ASH- or SSH-like lesions on the background of chronic liver injury. CONCLUSION: Our data indicated that the cut-off period between ASH and SSH is in accordance with the Scheme of Viral Hepatitis Prevention and Therapy, China, published in 2000, but excluded a part of child SH cases. In our study, the authors found a few pathological features in ASH and SSH.