[Survival analysis of unexpected small cell lung cancer following surgery].

Objective: To investigate the survival and influencing factors of unexpected small cell lung cancer following surgery. Methods: We respectively reviewed the clinical characters of 104 patients who underwent surgical treatment and be proved as small cell lung cancer by pathology between January 2000 to October 2020 in Chinese PLA General Hospital. Overall survival (OS) of patients was evaluated using Kaplan-Meier and Cox proportional hazards analysis. Results: Of 104 patients, 27 cases showed central lesions, and other 77 showed peripheral nodules. The margin of nodules was smooth in 42 cases on CT imaging. The median OS was 34.3 months and 5-year OS rate was 45.8%. Postoperative 5-year OS rates for patients were 52.1%, 45.4%, and 27.8% for clinical stages Ⅰ, Ⅱ, and Ⅲ, respectively. Univariate analyses identified the age, surgical access, surgical approach, N stage, TNM stage and vascular cancer emboli were associated with OS (P<0.05). The N stage was an independent factor for the OS of patients (P<0.05). Conclusions: Patients with unexpected SCLC, including Ⅰ, Ⅱ and part ⅢA stage have favorable outcome and can benefit from surgery and systemic postoperative treatment. Standard lobectomy plus systemic lymph node dissection is commended.

[Recommendations of diagnosis and treatment of polyarteritis nodosa].

Polyarteritis nodosa (PAN) is a rare vasculitis that mainly involves small and medium arteries. It often occurs at the points where the vessels bifurcate, leading to microaneurysm formation, thrombosis, aneurysm rupture and bleeding, and infarction of organs.About a third of cases are associated with hepatitis B virus (HBV) infection.All tissues and organs of the body can be affected, with skin, joints and peripheral nerves being the most common.The pathological changes were fibrinoid necrosis, inflammatory cell infiltration and luminal thrombosis in the acute stage, and fibrous hyperplasia in the chronic stage.Overall outcomes for the disease have improved in recent decades, mainly reflecting early diagnosis and more effective treatments.The main treatments for PAN are glucocorticoid and cyclophosphamide.Patients with HBV-associated PAN should receive antiviral therapy and plasma exchange.

[Clinical cure strategies for hepatitis B: direct-acting antiviral drugs].

Direct-acting antivirals (DAAs) play a critical role for the therapy of chronical hepatitis B. DAAs can decrease the production of viral progeny of hepatitis B virus (HBV), breaking the viral dynamic equilibrium between: (1) virion production from hepatocytes and clearance from circulation; (2) replenishment and decay of covalently closed circular (ccc)DNA pool inside infected hepatocytes. Nucleos(t)ide analogues can potently shift the first balance to undetectable viremia in the blood, but have limited or no effect on the second one, thus making it imperative to develop new agents targeting additional step(s) of HBV life cycle. We herein briefly introduce the DAAs currently in development by classifying them as agents affecting the replenishment or the decay of cccDNA pool.

Expression of Ras-related protein 25 predicts chemotherapy resistance and prognosis in advanced non-small cell lung cancer.

Ras-related protein 25 (Rab25) is involved in many human malignancies. However, its role in chemotherapy response and prognosis in advanced non-small cell lung cancer (NSCLC) remains unknown. Therefore, we investigated the relationship between Rab25 and chemotherapy sensitivity and prognosis in NSCLC. Rab25 expression was assessed using immunohistochemistry in 324 advanced NSCLC patients. Its correlations with clinical features were analyzed. Sensitivity to cisplatin (DDP) was compared between DDP-sensitive A549 and DDP-resistant A549/DDP cells. Furthermore, small interfering RNA (siRNA) targeting Rab25 was used for in vitro experiments. Patients with positive Rab25 expression had a significantly lower chemotherapy response rate (P = 0.004) and poorer overall survival (OS, P = 0.0012) than those with negative Rab25 expression. Multivariate analysis indicated that Rab25 expression was an independent prognostic factor for OS (P = 0.016). Moreover, Rab25 expression was significantly higher in A549/DDP cells than in A549 cells. Knockdown of Rab25 by siRNA suppressed cell migration and invasion. Cisplatin resistance in A549/DDP cells was also partially reversed by Rab25 silencing. Rab25 expression is a potential prognostic index for advanced NSCLC patients and its inhibition may improve chemosensitization in NSCLC treatment.

Knockdown of NEAT1 mitigates ox-LDL-induced injury in human umbilical vein endothelial cells via miR-30c-5p/TCF7 axis.

OBJECTIVE: Atherosclerosis is an inflammation-associated disease resulting in a huge health hazard. Abundance of researches showed that long non-coding RNAs (lncRNAs) played vital roles in atherosclerosis, but the molecular mechanism of nuclear-enriched abundant transcript (NEAT1) has not been fully elucidated yet. PATIENTS AND METHODS: Human umbilical vein endothelial cells (HUVECs) were treated with oxidized low-density lipoprotein (ox-LDL) for constructing the model of atherosclerosis. The detection of NEAT1, microRNA-30c-5p (miR-30c-5p), and transcription factor 7 (TCF7) expression was implemented by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Cell proliferation and apoptosis were measured by 3-(4, 5-dimethylthiazol-2-y1)-2, 5-diphenyl tetrazolium bromide (MTT) and flow cytometry, respectively. The levels of apoptosis-associated proteins were examined through Western blot and the concentrations of inflammatory cytokines were determined by enzyme-linked immunosorbent assay (ELISA). The targeted relationship was analyzed by Dual-Luciferase reporter assay. RESULTS: NEAT1 was upregulated in serum of patients with atherosclerosis and HUVECs treated with ox-LDL. Knockdown of NEAT1 exerted the promotion of proliferation but suppression of apoptosis and inflammation in ox-LDL-treated HUVECs. Moreover, NEAT1 targeted miR-30c-5p and the overexpression of miR-30c-5p reversed the ox-LDL-induced effects in HUVECs. Furthermore, miR-30c-5p directly refrained the TCF7 level, and NEAT1 repression decreased the expression of TCF7 by upregulating miR-30c-5p. The knockdown of NEAT1 afforded the protective effect for HUVECs treated with ox-LDL through miR-30c-5p/TCF7 axis. CONCLUSIONS: The knockdown of NEAT1 overtly motivated proliferation but alleviated the apoptosis and inflammation in ox-LDL-treated HUVECs by miR-30c-5p/TCF7 axis. NEAT1 accelerated the progression of atherosclerosis therapies, functioning as an indicative element.

[Verification of clinical applicability of the non-special perioperative administration for enhanced recovery after surgery of gastric cancer patients: a Chinese single-center observational report].

Objective: To verify clinical applicability of the non-special perioperative administration for enhanced recovery after surgery (ERAS) proposed by Japanese scholars in Chinese gastric cancer patients. Methods: The main measures of the non-special perioperative administration for ERAS are as follows: (1) discussion of multiple disciplinary team before surgery; (2) rehabilitation education for patients; (3) no routine bowel preparation before surgery; (4) placement of nasogastric tube for decompression routinely before operation and removal as early as 24 hours after surgery; (5) appropriate rehydration; (6) antibiotic prophylaxis before surgery; (7) place abdominal drainage tubes when necessary; (8) epidural patient-controlled analgesia and oral medication for postoperative pain management; (9) start low-molecular-weight heparin injection 48h after surgery and ambulation every day to prevent deep vein thrombosis; (10) postoperative dietary management and supplement with parenteral nutrition intermittently; (11) remove Foley catheter about 24 hours after surgery. A retrospective cohort study was performed, including 203 patients undergoing radical gastrectomy at Department of Gastroenterology, Tianjin Medical University Cancer Institute and Hospital from January 2017 to December 2018. Inclusion criteria were patients who were ≤75 years old without distant metastasis by preoperative examination, were diagnosed as gastric adenocarcinoma by postoperative histopathology and had complete clinicopathological and follow-up data. Patients with history of other malignancies and gastrectomy, extensive implantation of the abdominal cavity or malignant ascites by intraoperative exploration, death within 1 month after surgery, and residual gastric cancer were excluded. The perioperative management methods were chosen by patients. There were 123 patients who followed non-special perioperative administration for ERAS (non-special preparation group) and 80 patients who underwent traditional perioperative management (traditional method group). The primary outcomes (postoperative hospital stay, time to the first flatus, time to the first fluid diet, time to the first ambulatory activity, morbidity of postoperative complication, mortality, and readmission rate) and secondary outcomes (operative time, intraoperative blood loss and postoperative pain score) were compared between the two groups. Results: Compared to the traditional method group, the non-special preparation group had shorter time to the first flatus [(3.6±1.1) days vs. (4.8±1.4) days, t=3.134, P=0.003], shorter time to the first liquid diet [(2.6±0.9) days vs. (5.5±1.6) days, t=15.105, P<0.001], shorter time to the first ambulatory activity [(1.9±0.5) days vs. (4.1±1.1) days, t=8.543, P<0.001] and shorter postoperative hospital stay [(9.6±2.3) days vs. (12.9±2.3) days, t=5.020, P<0.001]. Besides, incidences of pancreatic leakage [6.5% (8/123) vs. 16.3% (13/80), χ(2)=4.964, P=0.026], lymphatic leakage [1.6% (2/123) vs. 13.8% (11/80), χ(2)=11.887, P=0.001], peritoneal effusion [2.4% (3/123) vs. 10.0% (8/80), χ(2)=4.032, P=0.045], and gastroparesis [0.8% (1/123) vs. 7.5% (6/80), χ(2)=4.657, P=0.031] in the non-special preparation group were significantly lower. The overall morbidity of postoperative complications and incidences of pulmonary infection and intestinal adhesion were not significantly different between the two groups (all P>0.05). As for the secondary outcomes, compared to the traditional method group, the non-special preparation group had less intraoperative blood loss [(80.4±24.4) ml vs. (100.5±19.4) ml, t=3.134, P=0.003] and lower postoperative pain score [postoperative day 1: (4.4±0.3) vs. (5.3±0.8), t=2.504, P=0.037],while the difference in operative time was not significant (P>0.05). Conclusion: The non-special perioperative administration for ERAS proposed by Japanese scholars is effective and safe, which has certain clinical applicability and value for Chinese patients with gastric cancer.

Tuning electronic and optical properties of monolayer PdSe2 by introducing defects: first-principles calculations.

Based on the density functional theory, the electronic and optical properties of pristine monolayer PdSe2 with Pd or Se vacancy-defect are investigated. Our results show that the Se defect is energetically more favorable than that of Pd defect. The band gap reduces, and some new midgap states appear after the Pd or Se defects are introduced. In terms of the optical properties, the prominent anisotropic characters are remained. The obvious new peaks of the dielectric constant appear after introducing defects. The light absorption in the visible energy range expands based on the appearance of the midgap states induced by the Pd or Se defects. The changes of the refractive index and reflectivity are similar with those of the dielectric constants and the light absorption. The energy loss spectrum of the PdSe2 with Pd or Se defects is obviously different, which can be used to identify different defects in PdSe2. These findings provide effective strategies to tune electronic and optical properties of monolayer PdSe2 by introducing defects.

Effect of IL-1ß on apoptosis of synovial cells in rheumatoid arthritis rats via the NF-κB pathway.

OBJECTIVE: The aim of this study was to investigate the role of interleukin-1ß (IL-1ß) in the apoptosis of synovial cells in rheumatoid arthritis (RA) rats, and to explore the underlying mechanism. MATERIALS AND METHODS: The apoptosis of the synovial cells in RA rats in the IL-1ß group and the control group was analyzed by scoring under an electron microscope. The expressions of cleaved-poly (ADP-ribose) polymerase (PARP), PARP and anti-apoptosis gene products in synovial cells of IL-1ß treated RA rats were explored as well. Meanwhile, the expressions of B-cell lymphoma 2 (Bcl-2), Bcl-xL, and Active-Caspase3 in the synovial cells of RA rats with IL-1ß treatment were evaluated by the Western blotting. To further clarify the relationship between IL-1ß and the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway in the synovial cells of RA rats, the expressions of NF-κB regulated the gene products of matrix metalloproteinase-3 (MMP-3), MMP-9, cyclooxygenase-2 (Cox-2), and vascular endothelial growth factor (VEGF) in synovial cells of RA rats after that we investigated the treatment with IL-1ß (was investigated). In addition, the expression of NF-κB in the synovial cells of RA rats treated with IL-1ß was determined. RESULTS: The results showed that, compared with the control group, IL-1ß treatment significantly increased the number of apoptotic cells. This meant that IL-1ß treatment could promote the apoptosis of the synovial cells (p<0.05). IL-1ß treatment significantly promoted the expression level of cleaved-PARP (p<0.05). However, it remarkably reduced the expressions of Bcl-2 and Bcl-xL (p<0.05). Meanwhile, the level of the active-Caspase3 in the synovial cells of RA rats treated with IL-1ß was significantly enhanced (p<0.01). In comparison with the control group, the IL-1ß group exhibited significantly elevated expressions of NF-κB-regulated gene products in the synovial cells of RA rats (p<0.01). Besides, the positive markers of the activated NF-κB were detected in the synovial cells of RA rats in the IL-1ß group and the control group. The results demonstrated that they were mainly located in the nucleus of the IL-1ß group. CONCLUSIONS: IL-1ß can promote the apoptosis of the synovial cells in RA rats via the NF-κB pathway.

Cloning and expression of a cDNA encoding the large subunit of duck replication factor C.

A cDNA encoding an avian homologue of the large subunit of replication factor C (RFC-L) has been cloned from a duck liver cDNA expression library prepared in bacteriophage lambda. The full length cDNA encodes a protein with a predicted size of approximately 130 kDa, consistent with the size of the polypeptide detected in duck liver. The duck RFC-L amino acid sequence shares 66.4% and 68.4% identity with mouse and human RFC-L proteins, respectively. We identified a 4kb RFC-L mRNA expressed in most duck tissues.

Amyloid formation in the rat: adenoviral expression of mouse serum amyloid A proteins.

Serum amyloid A (SAA) proteins are acute-phase apolipoproteins that are associated with high-density lipoprotein (HDL) particles: SAA proteins are precursors to secondary amyloid fibril proteins and under certain conditions of chronic or recurrent inflammation these proteins are deposited as amyloid fibrils. Of two isotypes found in mouse, SAA1.1 and SAA2.1, only SAA1.1 is deposited into amyloid. The CE/J mouse is unique, in that the only isoform identified is a hybrid between SAA1.1 and SAA2.1 and the mouse does not show amyloid deposition. In the rat, a deletion in the SAA1/SAA2 gene is associated with the absence of protein in the plasma and subsequently no amyloid deposition is detected. We have generated adenoviral vectors to study the expression of SAA proteins on HDL metabolism and amyloid formation. Injection of SAA viruses into rats resulted in expression of the mouse SAA proteins in the plasma with specific association of the SAA with HDL particles. The induction of SAA proteins was comparable to that seen in mice presented with the inflammatory agent, bacterial lipopolysaccharide (LPS). Adenoviral induced SAA levels were maintained for up to several weeks without a significant decrease in SAA expression. Injection of rats with the mouse SAA1.1 adenoviral vector, followed by amyloid enhancing factor (AEF) and silver nitrate resulted in the deposition of amyloid fibrils in the spleen. After 2 weeks, amyloid could be detected in other tissues, including the heart, liver, kidneys and lungs. When animals were injected with null or the SAA2.2 virus no amyloid was detected. These studies demonstrate that the inability of the rat to develop AA amyloid is due to the lack of synthesizing an amyloidogenic SAA protein. Furthermore, the expression of the adenoviral SAA protein from the liver and incorporation onto HDL particles further supports the hypothesis that AA amyloid is derived from circulating SAA protein. The ease of use of the adenoviral vectors and the rat provide an excellent model to study the function of SAA proteins.

[Uses of duck hepatitis B virus polymerase and reverse transcriptase in the evaluation of anti-hepatitis B virus drugs].

The method of DHBV replication complexes (RCs) purification was modified. In order to screen anti-HBV drugs from Chinese medicinal herbs, the inhibitory effects of the extracts of 14 Chinese recipes and herbs, 30 compounds isolated from Chinese herbs on DHBV DNA polymerase (DNAP) and reverse transcriptase (RT) have been studied. The results showed that extracts of Xiao Chai Hu Tang (small Bupleurum decoction) inhibited DHBV DNAP and RT in less extent. Of the 7 herbs, the components of Xiao Chai Hu Tang, the extracts of S. baicalensis and P. ternata potently inhibited DHBV RT, their concentration of reducing enzyme activity by 50% (IC50) was 1.25 and 1.6 mg/ml respectively. Furthermore, it has been proved that S. baicalensis inhibited DHBV DNA replication in ducklings. It also was found the extract of P. cuspidatum inhibited DHBV RT with IC50 of 1.76 mg/ml. Nine of thirty isolated compounds inhibited both DHBV DNAP and RT in less extent under high concentration, while other did not.

Topology of the large envelope protein of duck hepatitis B virus suggests a mechanism for membrane translocation during particle morphogenesis.

We have investigated the membrane topology of the large envelope protein of duck hepatitis B virus (DHBV) by protease protection and Western blot analysis, using monoclonal antibodies specific for the pre-S and S regions of the DHBV envelope to characterize protease-resistant polypeptides. These studies showed that DHBV L protein exhibits a mixed membrane topology similar to that of human hepatitis B virus L, with approximately half of the L molecules displaying pre-S on the surface of virus particles and the remainder with pre-S sequestered inside the virus envelope. The C-terminal region of DHBV pre-S was susceptible to protease digestion on all DHBV particle L protein, indicating that this region was externally disposed. DHBV L protein pre-S was entirely cytosolic immediately after synthesis. Our data, therefore, suggested that an intermediate form of the DHBV L molecule exists in mature envelope particles in which L is partially translocated or exists in a translocation-ready conformation. Incubation of virus particles at low pH and 37 degrees C triggered conversion of this intermediate into a fully translocated form. We have proposed a model for pre-S translocation based on our results that invokes the presence of an aqueous pore in the virus envelope, most likely created by oligomerization of transmembrane domains in the S region. The model predicts that pre-S is transported through this pore and that a loop structure is formed because the N terminus remains anchored to the inner face of the membrane. This translocation process occurs during particle morphogenesis and may also be a prerequisite to virus uncoating during infection.

Monoclonal antibodies to a 55-kilodalton protein present in duck liver inhibit infection of primary duck hepatocytes with duck hepatitis B virus.

As an approach to identifying hepatocyte receptors for the avian hepadnavirus duck hepatitis B virus (DHBV), hybridomas were prepared from mice immunized with permissive duck hepatocytes. Monoclonal antibodies (MAbs) were screened for the ability to inhibit binding of DHBV particles to primary duck hepatocytes and to block infection. We identified two MAbs which partially blocked binding and caused marked inhibition of infection of primary duck hepatocytes with DHBV. Lack of cross-reactivity with DHBV envelope proteins suggested that inhibition of infection was due to specific interaction between the antibodies and a host cell surface molecule. Both MAbs immunoprecipitated a 55-kDa protein (p55) expressed in duck liver and several other duck tissues. p55 homologs were also identified in other birds and mammals. We predict from our data that only a small proportion of total cellular p55 molecules are expressed at the surfaces of hepatocytes and that p55 is involved in some early step in the infectious pathway.

Protein fold recognition through application of residual dipolar coupling data.

Residual dipolar coupling (RDC) represents one of the most exciting emerging NMR techniques for studying protein structures. However, solving a protein structure using RDC data alone is a highly challenging problem as it often requires that the starting structure model be close to the actual structure of a protein, for the structure calculation procedure to be effective. We report in this paper a computer program, RDC-PROSPECT, for identification of a structural homolog or analog of a target protein in PDB, which best matches the 15N-1H RDC data of the protein recorded in a single ordering medium. The identified structural homolog/analog can then be used as a starting model for RDC-based structure calculation. Since RDC-PROSPECT uses only RDC data and predicted secondary structure information, its performance is virtually independent of sequence similarity between a target protein and its structural homolog/analog, making it applicable to protein targets out of the scope of current protein threading techniques. We have tested RDC-PROSPECT on all 15N-1H RDC data (representing 33 proteins) available in the BMRB database and the literature. The program correctly identified the structural folds for approximately 80% of the target proteins, significantly better than previously reported results, and achieved an average alignment accuracy of 97.9% residues within 4-residue shift. Through a careful algorithmic design, RDC-PROSPECT is at least one order of magnitude faster than previously reported algorithms for principal alignment frame search, making our algorithm fast enough for large-scale applications.

Effect of alpha interferon on the hepatitis C virus replicon.

Chronic hepatitis C virus (HCV) infections can be cured only in a fraction of patients treated with alpha interferon (IFN-alpha) and ribavirin combination therapy. The mechanism of the IFN-alpha response against HCV is not understood, but evidence for a role for viral nonstructural protein 5A (NS5A) in IFN resistance has been provided. To elucidate the mechanism by which NS5A and possibly other viral proteins inhibit the cellular antiviral program, we have constructed a subgenomic replicon from a known infectious HCV clone and demonstrated that it has an approximately 1,000-fold-higher transduction efficiency than previously used subgenomes. We found that IFN-alpha reduced replication of HCV subgenomic replicons approximately 10-fold. The estimated half-life of viral RNA in the presence of the cytokine was about 12 h. HCV replication was sensitive to IFN-alpha independently of whether the replicon expressed an NS5A protein associated with sensitivity or resistance to the cytokine. Furthermore, our results indicated that HCV replicons can persist in Huh7 cells in the presence of high concentrations of IFN-alpha. Finally, under our conditions, selection for IFN-alpha-resistant variants did not occur.

Expression of mouse apolipoprotein SAA1.1 in CE/J mice: isoform-specific effects on amyloidogenesis.

Amyloid A (AA) amyloid deposition in mice is dependent upon isoform-specific effects of the serum amyloid A (SAA) protein. In type A mice, SAA1.1 and SAA2.1 are the major apolipoprotein-SAA isoforms found on high-density lipoproteins. During inflammation, both isoforms are increased 1000-fold, but only SAA1.1 is selectively deposited into amyloid fibrils. Previous studies showed that the CE/J mouse strain is resistant to amyloid induction. This resistance is not due to a deficiency in SAA synthesis, but is probably related to the unusual SAA isoform present. The CE/J mouse has a single acute-phase SAA protein (SAA2.2), which is a composite of the SAA1.1 and SAA2.1, with an amino terminus similar to the nonamyloidogenic SAA2.1. Recently, genetic experiments suggested that the SAA2.2 isoform might provide protection from amyloid deposition. To determine the amyloidogenic potential of the CE/J mouse, we generated SAA adenoviral vectors to express the various isoforms in vitro and in vivo. Purified recombinant SAA proteins demonstrated that SAA1.1 was fibrillogenic in vitro, whereas SAA2.2 was unable to form fibrils. Incubation of increasing concentrations of the nonamyloidogenic SAA2.2 protein with the amyloidogenic SAA1.1 did not inhibit the fibrillogenic nature of SAA1.1, or alter its ability to form extensive fibrils. Injection of the mouse SAA1.1 or SAA2.2 adenoviral vectors into mice resulted in isoform-specific expression of the SAA proteins. Amyloid induction after viral expression of the SAA1.1 protein resulted in the deposition of amyloid fibrils in the CE/J mouse, whereas SAA2.2 expression had no effect. Similar expression of the SAA2.2 protein in C57BL/6 mice did not alter amyloid deposition. These data demonstrate that the failure of the CE/J mouse to deposit amyloid is due to the structural inability of the SAA2.2 to form amyloid fibrils. This mouse provides a unique system to test the amyloidogenic potential of altered SAA proteins and to determine the important structural features of the protein.

Does a cdc2 kinase-like recognition motif on the core protein of hepadnaviruses regulate assembly and disintegration of capsids?

Hepadnaviruses are enveloped viruses, each with a DNA genome packaged in an icosahedral nucleocapsid, which is the site of viral DNA synthesis. In the presence of envelope proteins, DNA-containing nucleocapsids are assembled into virions and secreted, but in the absence of these proteins, nucleocapsids deliver viral DNA into the cell nucleus. Presumably, this step is identical to the delivery of viral DNA during the initiation of an infection. Unfortunately, the mechanisms triggering the disintegration of subviral core particles and delivery of viral DNA into the nucleus are not yet understood. We now report the identification of a sequence motif resembling a serine- or threonine-proline kinase recognition site in the core protein at a location that is required for the assembly of core polypeptides into capsids. Using duck hepatitis B virus, we demonstrated that mutations at this sequence motif can have profound consequences for RNA packaging, DNA replication, and core protein stability. Furthermore, we found a mutant with a conditional phenotype that depended on the cell type used for virus replication. Our results support the hypothesis predicting that this motif plays a role in assembly and disassembly of viral capsids.

Characterization of serum amyloid A protein mRNA expression and secondary amyloidosis in the domestic duck.

Secondary amyloidosis is a common disease of water fowl and is characterized by the deposition of extracellular fibrils of amyloid A (AA) protein in the liver and certain other organs. Neither the normal role of serum amyloid A (SAA), a major acute phase response protein, nor the causes of secondary amyloidosis are well understood. To investigate a possible genetic contribution to disease susceptibility, we cloned and sequenced SAA cDNA derived from livers of domestic ducks. This revealed that the three C-terminal amino acids of SAA are removed during conversion to insoluble AA fibrils. Analysis of SAA cDNA sequences from several animals identified a distinct genetic dimorphism that may be relevant to susceptibility to secondary amyloid disease. The duck genome contained a single copy of the SAA gene that was expressed in liver and lung tissue of ducklings, even in the absence of induction of acute phase response. Genetic analysis of heterozygotes indicated that only one SAA allele is expressed in livers of adult birds. Immunofluorescence staining of livers from adult ducks displaying early symptoms of amyloidosis revealed what appear to be amyloid deposits within hepatocytes that are expressing unusually high amounts of SAA protein. This observation suggests that intracellular deposition of AA may represent an early event during development of secondary amyloidosis in older birds.

Increased serum amyloid a levels reflect colitis severity and precede amyloid formation in IL-2 knockout mice.

The lack of sensitive and relatively non-invasive measures has hampered monitoring the clinical course of spontaneously developing colitis in IL-2-deficient (-/-) mice. We selected (i) to study the correlation of the acute phase plasma proteins serum amyloid A (SAA) and serum amyloid P component (SAP) levels with colonic disease and (ii) to characterize the amyloidosis in the IL-2(-/-)animals. IL-2(-/-)mice exhibited increasing severity of gross intestinal inflammation with age, confined to the distal colon. Histologically, the colonic disease score increased serially in IL-2(-/-)animals. Wild-type mice showed no activity, while 16-week-old IL-2(+/-)animals had minimal colitis with small ulcers and lamina propria inflammatory infiltrate. Periportal hepatitis was present and positive Congo red staining indicated amyloidosis of the liver and spleen in 16 week IL-2(-/-)mice. SAA immunostaining in the liver and spleen was increased in the 8 week and 16 week IL-2(-/-)and 16 week IL-2(+/-)animals indicating AA amyloid deposits. Plasma SAA and SAP levels were markedly elevated, and generally preceded the onset of colitis and reflected its severity. Northern analysis showed markedly increased SAA expression in the liver and intestine of IL-2(-/-)and intestine of IL-2(+/-)16-week-old animals. Increased intestinal expression of SAA3 (lamina propria macrophages) indicates local inflammation in IL-2(+/-)animals at 16 weeks. Treatment of 3-week-old animals with systemic IL-2 or IL-1 receptor antagonist (IL-1ra) delayed inflammation, postponed the increase in SAA levels and minimized disease onset. These results further demonstrate that IL-2 plays a significant role in normal immune responses in the body and that plasma SAA levels both reflect colonic disease severity and may indicate subclinical disease in both IL-2(-/-)and IL-2(+/-)mice. Furthermore. The mechanism of IL-2-deficient induced colitis appears to be mediated in part through the increase in IL-1. In addition, the IL-2(-/-)mouse of spontaneous enterocolitis may provide a unique system for studying spontaneously developing AA amyloidosis.