[Curative effect of percutaneous microwave ablation therapy on hepatocellular carcinoma survival: a 15-year real-world study].

Objective: To evaluate the long-term efficacy of percutaneous microwave ablation (MWA) therapy for hepatocellular carcinoma. Methods: 2054 cases with Barcelona Clinic Liver Cancer (BCLC) stage 0~B at the Fifth Medical Center of the Chinese People's Liberation Army General Hospital from January 2006 to September 2020 were retrospectively collected. All patients were followed up for at least 2 years. The primary endpoint of overall survival and secondary endpoints (tumor-related survival, disease-free survival, and postoperative complications) of patients treated with ultrasound-guided percutaneous MWA were analyzed. Kaplan-Meier method was used for stratified survival rate analysis. Fine-and-Gray competing risk model was used to analyze overall survival. Results: A total of 5 503 HCC nodules [mean tumor diameter (2.6±1.6) cm] underwent 3 908 MWAs between January 2006 and September 2020, with a median follow-up time of 45.6 (24.0 -79.2) months.The technical effectiveness rate of 5 375 tumor nodules was 97.5%. The overall survival rates at 5, 10, and 15-years were 61.6%, 38.8%, and 27.0%, respectively. The tumor-specific survival rates were 67.1%, 47.2%, and 37.7%, respectively. The free tumor survival rates were 25.8%, 15.7%, and 9.9%, respectively. The incidence rate of severe complications was 2.8% (108/3 908). Further analysis showed that the technical effectiveness and survival rate over the passing three time periods from January 2006-2010, 2011-2015, and 2016-September 2020 were significantly increased, with P < 0.001, especially for liver cancer 3.1~5.0 cm (P < 0.001). Conclusion: Microwave ablation therapy is a safe and effective method for BCLC stage 0-B, with significantly enhanced technical efficacy and survival rate over time.

Cloning and expression analysis of cysteine protease gene (MwCP) in Agropyron mongolicum Keng.

In this study, a cysteine protease gene (MwCP) from Agropyron mongolicum Keng was isolated using RACE. Sequence analysis indicated that MwCP was 1473 bp, and it contained a 1134-bp open reading frame, which encoded 377 amino acids with a 24-amino acid N-terminal signal peptide. The results indicated that the MwCP protein was a new member of the papain C1A family, and it was predicted to be an extracellular, secretory stable hydrophilic protein. The secondary structure of MwCP was mainly composed of α-helices and random coils, and the space structure primarily contained α-helices, ß-sheets, and ß-turns. Homology analyses showed the 98% homology between MwCP amino acids and a cysteine protease found in Triticum aestivum (GenBank accession No. AAW21813.1). Analysis of mRNA using semi-quantitative RT-PCR indicated that during a 48-h drought stress period, MwCP was expressed during the 4th hour, and the expression level peaked during the 6th hour before declining to the original level. The results revealed that MwCP was involved in drought-resistant physiological processes of A. mongolicum. Moreover, the MwCP expression levels were highest in leaves, intermediate in roots, and lowest in stems.

Fc alpha receptor cross-linking causes translocation of phosphatidylinositol-dependent protein kinase 1 and protein kinase B alpha to MHC class II peptide-loading-like compartments.

A20 IIA1.6 B cells cotransfected with FcalphaR and wild-type gamma-chain (wt-ITAM (immunoreceptor tyrosine-based activation motif)) or FcalphaR and gamma-chain, in which the wt-ITAM was substituted with the FcgammaRIIA ITAM (IIA-ITAM), were used to investigate cell signaling events influencing presentation of FcalphaR-targeted exogenous Ag in the context of MHC class II. wt-ITAM cells presented FcalphaR-targeted OVA more efficiently than IIA-ITAM transfectants to OVA-specific T cell hybridomas. Phosphatidylinositol 3-kinase (PI 3-kinase) inhibition abrogated Ag presentation, suggesting that FcalphaR may trigger a PI 3-kinase-dependent signal transduction pathway, and thus phosphatidylinositol-dependent protein kinase (PDK1) and protein kinase B alpha (PKBalpha) activation. Cross-linking FcalphaR on wt-ITAM or IIA-ITAM cells triggered equivalent PI 3-kinase-dependent activation of PKBalpha. Furthermore, FcalphaR cross-linking triggered recruitment of PDK1 and serine-phosphorylated PKBalpha to capped cell surface FcalphaR irrespective of the gamma-chain ITAM. Although FcalphaR endocytosis was accompanied by translocation of PDK1 and phospho-PKBalpha to FcalphaR-containing vesicles in both transfectants, this was decreased in IIA-ITAM cells, and a significant proportion of PDK1 and PKBalpha remained at the plasma membrane. In wt-ITAM cells, PDK1 and serine-phosphorylated PKBalpha translocated to lysosomal-associated membrane glycoprotein 1- and cathepsin B-containing vesicles, consistent with MHC class II peptide-loading compartments (MIIC) described by other groups. Our data indicate that translocation of signal transduction mediators to MIIC-like compartments accompanies efficient presentation of receptor-targeted Ag, and suggest a mechanism connecting signaling to the Ag-processing pathway.

Presentation of ovalbumin internalized via the immunoglobulin-A Fc receptor is enhanced through Fc receptor gamma-chain signaling.

The mechanism of enhanced presentation of ovalbumin (OVA) internalized as immunoglobulin A (IgA)-OVA via the IgA Fc receptor (FcalphaR) was analyzed by focusing on the role of the FcalphaR-associated gamma chain. Comparison of B-cell transfectants expressing FcalphaR plus wild-type (WT) gamma chain or gamma chain in which the immunoreceptor tyrosine-based activation motif (ITAM) was altered by tyrosine mutation or substitution with the ITAM of FcgammaRIIA showed that signaling-competent ITAM was not required for endocytosis of IgA-OVA. However, antigen presentation was impaired by ITAM changes. Signaling-competent gamma-chain ITAM appeared necessary for transport of ligated FcalphaR to a lamp-1(+) late endocytic compartment for remodeling and/or activation of that compartment and also for efficient degradation of IgA complexes. Moreover, FcalphaR ligation also activated efficient processing of nonreceptor-targeted antigen. The results suggest that gamma-chain signaling activates the antigen processing compartment.

Characterization of FcalphaR-triggered Ca(2+) signals: role in neutrophil NADPH oxidase activation.

Human neutrophil IgA receptors (FcalphaR) trigger phagocytosis of IgA-opsonized particles and activate the NADPH oxidase complex ultimately leading to pathogen destruction. Signal transduction events triggered by FcalphaR have not been investigated in the context of NADPH oxidase activation. In this study, we show that crosslinking FcalphaR triggers the release of Ca(2+) from an intracellular store that was unchanged by the addition of extracellular EGTA. This was in contrast to the thapsigargin-triggered Ca(2+) signal, which activates store-operated Ca(2+) entry pathways (SOCP) and is sensitive to extracellular EGTA. Buffering extracellular Ca(2+) with EGTA had no effect on FcalphaR-triggered NADPH oxidase activation, suggesting that SOCP was not required for activation by FcalphaR. EGTA inhibited thapsigargin-triggered NADPH oxidase activation but had no effect on PMA-triggered responses. The intracellular Ca(2+) chelator BAPTA caused dose-dependent inhibition of both FcalphaR-triggered and thapsigargin-triggered NADPH oxidase activation but had no effect on PMA-triggered responses. Our data demonstrate that FcalphaR-triggered NADPH oxidase activation is dependent on the release of Ca(2+) from an intracellular store, but is independent of SOCP.

The ins and outs of getting in: structures and signals that enhance BCR or Fc receptor-mediated antigen presentation.

Antigen-presenting cells internalize antigen by fluid-phase pinocytosis or by endocytosis via surface receptors such as the B cell receptor (BCR) and Fc receptors for IgG, IgA and IgE (FcR). While both modes of internalization lead to antigen presentation it is recognized that receptor-mediated endocytosis greatly enhances the efficiency of processing and antigen presentation. Receptors facilitate the entry of antigen into the endocytic pathway by interaction of their internalization motifs with the endocytic machinery. These motifs include tyrosine-based, dileucine and casein kinase-like motifs. However these structures appear insufficient to support processing of cryptic epitopes, leading to a limited immune response. Cryptic epitope processing appears dependent on receptor signaling which is mediated by immunoreceptor tyrosine activation motifs (ITAMs). The signaling cascade which follows receptor crosslinking promotes reorganization and acidification of the late endocytic compartment or MIIC. Signaling events downstream of Syk, in particular calcium flux and protein kinase C activation, are necessary for MIIC induction. PI(3) kinase is also involved at multiple steps in antigen presentation, including production of PIP3 and transport of cathepsins. PIP3 is crucial both as a binding substrate for proteins implicated in vesicle transport and for the recruitment of signaling molecules to the plasma membrane. Among PIP3 activated molecules, protein kinase B (PKB) has been linked to endocytic function. We observe association of activated PKB with the MIIC after signaling through antigen presentation-competent receptors, but not mutant, presentation-defective receptors.

Mutations in specific I-A(k) alpha(2) and beta(2) domain residues affect surface expression.

A previous investigation demonstrated that several mutations in class II dimer-of-dimers contact residues interfere with antigen presentation by transfectants but not with plasma membrane expression of the mutant class II. In the present study we examined other class II mutations in this region that did inhibit plasma membrane expression of mutant class II molecules. Molecules containing both mutations H alpha 181D in the alpha(2) domain and E beta 170K in the beta(2) domain exhibited low plasma membrane expression, but molecules with only one of these mutations were expressed normally. The mutant class II molecules were transported to organelles that were accessible to a fluid-phase protein, hen egg lysozyme (HEL). Culture of transfectants with lysozyme enhanced the amount of class II compact dimer (alpha beta plus peptide; CD), and this was especially marked for the class II mutant H alpha 181D/E beta 170K and for other molecules possessing both mutations. Formation of class II CD was not paralleled by an increase in class II surface expression. Thus the joint mutation of H alpha 181 and E beta 170 has two effects. In the absence o high concentrations of exogenous peptide, it prevents efficient CD formation, possibly by affecting invariant chain (Ii) proteolysis and/or the stability of the class II after Ii/CLIP is removed. At high peptide concentrations supplied by exogenous HEL, the mutations allow CD formation, but not expression of class II on the plasma membrane. Molecular modeling of the possible interaction of class II and Ii suggests that the mutant amino acids H alpha 181D and E beta 170K, besides affecting the overall stability of class II, might also interact with Ii via two loops in class II's alpha(2) and beta(2) domains respectively.

Neutrophil NADPH oxidase does not assemble on macropinocytic vacuole membranes.

NADPH oxidase assembly is considered to occur mainly on the plasma membrane and phagolysosome membranes and to a lesser extent in intracellular granules. Stimulation of neutrophils with phorbol 12-myristate 13-acetate (PMA) in conjunction with chemiluminescent substrates has been widely used as a model to understand the sub-cellular locations of NADPH-oxidase activity. Interpretation of data from such studies is complicated by observations that PMA does not trigger phagocytosis but results in formation of large macropinocytic vacuoles. Here we show by laser-scanning confocal microscopy that PMA triggers uptake of lucigenin into macropinocytic vacuoles and that no chemiluminescence (CL) can be detected at this location when NADPH oxidase assembly on the plasma membrane still occurs. This shows that the macropinocytic vacuole membrane is distinct from the phagocytic vacuole membrane although both compartments are derived from the plasma membrane.

Gamma-chain dependent recruitment of tyrosine kinases to membrane rafts by the human IgA receptor Fc alpha R.

We show that the human IgA receptor, Fc alpha R, redistributes to plasma membrane rafts after cross-linking and that tyrosine kinases are relocated to these sites following Fc alpha R capping. We demonstrate by confocal microscopy that Fc alpha R caps in membrane rafts by a gamma-chain-independent mechanism but that gamma-chain expression is necessary for Lyn redistribution. Immunoblotting of rafts isolated by sucrose density gradient centrifugation demonstrated recruitment of gamma-chain and phosphorylated tyrosine kinases Lyn and Bruton's tyrosine kinase to membrane rafts after Fc alpha R cross-linking. Time-dependent differences in Lyn phosphorylation and Bruton's tyrosine kinase distribution were observed between cells expressing Fc alpha R plus gamma-chain and cells expressing Fc alpha R only. This study defines early Fc alpha R-triggered membrane dynamics that take place before Fc alpha R internalization.

Diversity and Longevity of the Soybean Debris Mycobiota in a No-Tillage System.

The survival of the mycobiota on pod and stem debris of soybean produced in a no-tillage system with cover crops of alfalfa, canola, rye, or wheat or with no cover was studied during 1994 and 1995. Fiberglass mesh bags containing pods and stems were assayed every 28 to 31 days to determine the isolation frequency of fungi. Over 90% of the 11,906 isolates obtained were members of the Deuteromycotina. The most common genera isolated were Alternaria, Cercos-pora, Colletotrichum, Epicoccum, Fusarium, and Phoma. Alternaria spp. had the greatest isolation frequencies and constituted 40% of the total cultures. Numbers of total fungi (all fungi isolated) on sampling dates in 1994 were similar to the totals in 1995. In May 1994, the mean isolation rates for many of the fungal species were significantly lower (P = 0.05) in several of the cover crops, but no consistent pattern could be determined. Common soybean pathogens isolated included Colletotrichum spp., Diaporthe spp., and Cercospora kikuchii. Fusarium graminearum, which is responsible for several diseases of maize and wheat, was commonly isolated during this study. Of the Diaporthe spp. (anamorph Phomopsis spp.), 87% were identified as D. phaseolorum var. sojae. Colletotrichum spp. were identified as C. truncatum in 85% of the isolates, C. destructivum (teleomorph Glomerella glycines) in 12%, and both species in 3%. Cercospora kikuchii was more commonly isolated from pods than from stem tissue, and Colletotrichum spp. occurred more frequently on stems. Isolation frequencies of Diaporthe spp. were greater in May of both years than in the preceding months. These results show that no-tillage soybean debris harbors numerous fungi pathogenic to soybean, and producers who grow soybeans continuously may find more disease in this crop and lower yields. Fungi that attack crops such as maize and wheat were commonly isolated from soybean debris in both years, and a no-tillage rotation which includes maize or wheat could result in increased disease in these crops. Isolation frequencies of the fungi from cover crops varied with the sampling date, but no consistent patterns could be determined for a particular cover crop or fungal species. This is the first detailed study of survival rates of soybean, maize, and wheat pathogens that overwinter on soybean debris in a no-tillage system.

Comparison of spinal and epidural anesthesia for patients younger than 50 years of age.

To evaluate the respective clinical value of spinal anesthesia with 24-gauge Sprotte needles and epidural anesthesia for younger subjects, 202 patients younger than 50 yr were assigned randomly to undergo one of these two techniques for orthopedic, vascular, urologic, or plastic surgery. Failed blocks occurred in 5% in each group. Spinal anesthesia resulted in significantly less time to achieve sufficient spread of block; a significantly lower incidence of incomplete sensory block at level L5/S1, incomplete motor block, and pain during surgery; and a significantly lower incidence of postlumbar puncture backache (11% vs 30% after epidural anesthesia). The incidence of postdural puncture headache (PDPH) in the spinal and epidural groups was 7% and 4%, respectively (P = not significant), and patient satisfaction was 97% and 93% (P = not significant). Our results demonstrate the effectiveness of both techniques in younger patients, but show that the spinal technique is associated with fewer limitations, suggesting that factors other than PDPH should be considered when choosing between these two techniques.