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OBJECTIVE: To explore the technical details and short-term effects of radiofrequency obliteration of varicose veins of lower extremities guided by combined venography and ultrasound. METHODS: Thirty-seven patients with varicose veins of lower extremities were treated with radiofrequency obliteration using Olympus Celon RFiTT(®) under combined guidance of venography and ultrasound. The indications included varicose veins of lower extremities and reflux of the great saphenous vein confirmed by ultrasound. The contraindications included deep vein thrombosis, cardiac pacemaker, severe cardio- and cerebrovascular diseases or coagulation disorders. Under ultrasound guidance, the saphenous vein around knee level was punctured using a 21G needle, and a 7F sheath was introduced. Through the sheath a venography was made, and an Olympus Celon ProCurve radiofrequency catheter was inserted and advanced to the great saphenous vein under road map, and the catheter tip was positioned at the point 2 cm below the sapheno-femoral junction. The swelling anesthesia was made under ultrasound guidance. Then the radiofrequency obliteration was performed with pressing of the treatment section. The venography was repeated to ensure optimal outcomes. If necessary the radiofrequency obliteration could be repeated once to twice. After that the superficial varicose veins were stripping by small incisions under local anesthesia. After operation, medical decompression stocking was utilized immediately and sustained for three months. The clinical data, intraoperative radiation dose, exposure time and short-term effects were retrospectively analyzed. RESULTS: After the operation, all the patients walked out of the operating room by themselves. The success rate of operation was 100%. The intraoperative radiation dose was 1.78-10.12 mGy (mean 6.56 mGy), and the exposure time was 61-448 s (mean 161 s). By 3 months follow-up, the symptoms were alleviated in all the 37 patients, and the occlusion rate was 100%. No complications such as skin burns, ecchymosis and deep venous thrombosis were found. CONCLUSION: The short-term effects of radiofrequency obliteration using Olympus Celon RFiTT(®) system in a manner of twice fixed point followed by once reciprocating radiofrequency were satisfactory. Radiofrequency obliteration of great saphenous veins guided by venography and ultrasound has not only the advantages of minimal trauma and rapid recovery, but also the advantages of accurate location, exact effect and avoidance of complications.

To uncover the potential function of LINC00628 in influencing the progression of colorectal cancer (CRC) and its underlying mechanism.Relative levels of LINC00628 and p57 in CRC tissues and cell lines were determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Regulatory effects of LINC00628 and p57 on proliferation, cell cycle, and apoptosis of SW480 and SW620 cells were assessed. Subcellular distribution of LINC00628 in CRC cells was analyzed. Moreover, the interaction between LINC00628 and enhancer of zeste homolog 2 (EZH2) was evaluated by the RNA Binding Protein Immunoprecipitation (RIP) assay.LINC00628 was downregulated in CRC tissues and cell lines. CRC patients expressing a low level of LINC00628 suffered worse prognosis. The. knockdown of LINC00628 enhanced proliferative ability, prolonged S phase in cell cycle progression, and inhibited apoptosis in SW480 and SW620 cells. LINC00628 was mainly distributed in the nucleus. The RIP assay demonstrated that LINC00628 could bind to EZH2 to upregulate the p57 level. Rescue experiments verified that the overexpression of p57 could reverse regulatory effects of downregulated LINC00628 on proliferative and apoptotic abilities of CRC.LINC00628 is downregulated in CRC. It aggravates the progression of CRC by binding to EZH2 to further inhibit the p57 level.

Summary During chronic inflammation, interleukin (IL)-22 expression is up-regulated in both CD4 and CD8 T cells, exerting a protective role in infections. However, in autoimmunity, IL-22 appears to have either a protective or a pathogenic role in a variety of murine models of autoimmunity and, by extrapolation, in humans. It is not clear whether IL-22 itself mediates inflammation or is a by-product of inflammation. We have taken advantage of the dominant negative form of transforming growth factor beta receptor type II (dnTGF-βRII) mice that develop both inflammatory bowel disease and autoimmune cholangitis and studied the role and the biological function of IL-22 by generating IL-22–/– dnTGF-βRII mice. Our data suggest that the influence of IL-22 on autoimmunity is determined in part by the local microenvironment. In particular, IL-22 deficiency exacerbates tissue injury in inflammatory bowel disease, but has no influence on either the hepatocytes or cholangiocytes in the same model. These data take on particular significance in the previously defined effects of IL-17A, IL-12p40 and IL-23p19 deficiency and emphasize that, in colitis, there is a dominant role of IL-23/T helper type 17 (Th17) signalling. Furthermore, the levels of IL-22 are IL-23-dependent. The use of cytokine therapy in patients with autoimmune disease has significant potential, but must take into account the overlapping and often promiscuous effects that can theoretically exacerbate inflammation.

Summary Cytotoxic T lymphocyte antigen 4 (CTLA-4) immunoglobulin (Ig) is an important regulator of T cell activation and a fusion protein directed at CD80 and CD86; it blocks co-stimulatory signalling and T cell activation. We have taken advantage of a murine model of human primary biliary cirrhosis (PBC), mice expressing a transforming growth factor (TGF)-β receptor II dominant negative (dnTGF-βRII) transgene to address the potential therapeutic efficacy of CTLA-4 Ig. To mimic patients with PBC at different stages or duration of disease, we treated mice with either CTLA-4 Ig or control IgG three times weekly from 3 to 12 or 24 weeks of age, or from 12 to 24 weeks of age. CTLA-4 Ig treatment from 3 weeks of age significantly reduced liver inflammation to 12 weeks of age. Treatment initiated at 12 weeks of age also ameliorated the autoimmune cholangitis at 24 weeks of age. However, in mice treated at 3 weeks of age, suppression of liver inflammation was not sustained and colitis was aggravated when treatment was extended to 24 weeks of age. Our data indicate that, in dnTGF-βRII mice, CTLA-4 Ig treatment has short-term beneficial effects on autoimmune cholangitis, but the effect varies according to duration of treatment and the time in which therapy was initiated. Further dissection of the events that lead to the reduction in therapeutic effectiveness of CTLA-4 Ig will be critical to determining whether such efforts can be applied to human PBC.

Summary Treatment of primary biliary cirrhosis (PBC) has lagged behind that of other autoimmune diseases. In this study we have addressed the potential utility of immunotherapy using regulatory T cells (Treg) to treat murine autoimmune cholangitis. In particular, we have taken advantage of our ability to produce portal inflammation and bile duct cell loss by transfer of CD8+T cells from the dominant negative form of transforming growth factor beta receptor type II (dnTGF-βRII) mice to recombination-activating gene (Rag)1–/– recipients. We then used this robust established adoptive transfer system and co-transferred CD8+T cells from dnTGF-βRII mice with either C57BL/6 or dnTGF-βRII forkhead box protein 3 (FoxP3+) T cells. Recipient mice were monitored for histology, including portal inflammation and intralobular biliary cell damage, and also included a study of the phenotypical changes in recipient lymphoid populations and local and systemic cytokine production. Importantly, we report herein that adoptive transfer of Treg from C57BL/6 but not dnTGF-βRII mice significantly reduced the pathology of autoimmune cholangitis, including decreased portal inflammation and bile duct damage as well as down-regulation of the secondary inflammatory response. Further, to define the mechanism of action that explains the differential ability of C57BL/6 Tregversus dnTGF-βRII Treg on the ability to down-regulate autoimmune cholangitis, we noted significant differential expression of glycoprotein A repetitions predominant (GARP), CD73, CD101 and CD103 and a functionally significant increase in interleukin (IL)-10 in Treg from C57BL/6 compared to dnTGF-βRII mice. Our data reflect the therapeutic potential of wild-type CD4+FoxP3+Treg in reducing the excessive T cell responses of autoimmune cholangitis, which has significance for the potential immunotherapy of PBC.

Summary While there have been significant advances in our understanding of the autoimmune responses and the molecular nature of the target autoantigens in primary biliary cirrhosis (PBC), unfortunately these data have yet to be translated into new therapeutic agents. We have taken advantage of a unique murine model of autoimmune cholangitis in which mice expressing a dominant negative form of transforming growth factor β receptor II (dnTGFβRII), under the control of the CD4 promoter, develop an intense autoimmune cholangitis associated with serological features similar to human PBC. CD40-CD40 ligand (CD40L) is a major receptor–ligand pair that provides key signals between cells of the adaptive immune system, prompting us to determine the therapeutic potential of treating autoimmune cholangitis with anti-CD40L antibody (anti-CD40L; MR-1). Four-week-old dnTGFβRII mice were injected intraperitoneally with either anti-CD40L or control immunoglobulin (Ig)G at days 0, 2, 4 and 7 and then weekly until 12 or 24 weeks of age and monitored for the progress of serological and histological features of PBC, including rigorous definition of liver cellular infiltrates and cytokine production. Administration of anti-CD40L reduced liver inflammation significantly to 12 weeks of age. In addition, anti-CD40L initially lowered the levels of anti-mitochondrial autoantibodies (AMA), but these reductions were not sustained. These data indicate that anti-CD40L delays autoimmune cholangitis, but the effect wanes over time. Further dissection of the mechanisms involved, and defining the events that lead to the reduction in therapeutic effectiveness will be critical to determining whether such efforts can be applied to PBC.

Summary The phagocytic clearance of apoptotic cells is critical for tissue homeostasis; a number of non-professional phagocytic cells, including epithelial cells, can both take up and process apoptotic bodies, including the release of anti-inflammatory mediators. These observations are particularly important in the case of human intrahepatic biliary cells (HiBEC), because such cells are themselves a target of destruction in primary biliary cirrhosis, the human autoimmune disease. To address the apoptotic ability of HiBECs, we have focused on their ability to phagocytize apoptotic blebs from autologous HiBECs. In this study we report that HiBEC cells demonstrate phagocytic function from autologous HiBEC peers accompanied by up-regulation of the chemokines CCL2 [monocyte chemotactic protein-1 (MCP-1)] and CXCL8 [interleukin (IL)-8]. In particular, HiBEC cells express the phagocytosis-related receptor phosphatidylserine receptors (PSR), implying that HiBECs function through the ‘eat-me’ signal phosphatidylserine expressed by apoptotic cells. Indeed, although HiBEC cells acquire antigen-presenting cell (APC) function, they do not change the expression of classic APC function surface markers after engulfment of blebs, both with and without the presence of Toll-like receptor (TLR) stimulation. These results are important not only for understanding of the normal physiological function of HiBECs, but also explain the inflammatory potential and reduced clearance of HiBEC cells following the inflammatory cascade in primary biliary cirrhosis.

Summary Although the hallmark of primary biliary cirrhosis (PBC) is the presence of anti-mitochondrial antibodies (AMA), a significant number of patients have anti-nuclear antibodies (ANA) directed primarily against two nuclear proteins, gp210 and sp100. In PBC, there are considerable data on the specificity of these anti-nuclear antibodies as well as suggestive evidence that antibodies to gp210 predict a poor outcome. However, a further understanding of the significance of these autoantibodies has been hampered by limitations in accessing human subjects in a preclinical or early asymptomatic stage. To overcome this limitation, we have taken advantage of transgenic mice with abrogated transforming growth factor-β signalling in T cells (dnTGF-βRII) that develop histological features of PBC as well as the same AMA specificity. We studied these mice for serum ANA, including specific autoantibodies against gp210 and sp100. We further examined sera from dnTGF-βRII mice with concurrent deletions of the genes encoding interleukin (IL)-12p35, IL-12p40, IL-23p19, IL-17, IL-6, interferon (IFN)-γ or tumour necrosis factor (TNF)-α. Sera from all the dnTGF-βRII mouse lines contained antibodies against gp210 and sp100. Of significance, mice with germline deletions of the genes encoding IL-12p40, IL-23p19, IL-17, IL-6 and TNF-α had significantly lower titres of anti-gp210 antibodies. These results provide a platform to dissect the mechanisms of gp210 and sp100 autoantibody production in dnTGF-βRII mice as well as to study the possible role of ANA in the pathophysiology of PBC.

The response of μ phase to applied stress and long-term thermal exposure has been investigated in the cast Ni-base superalloy K446. It is found that during stress rupture, the applied stress accelerates the precipitation and growth of μ phase. However, during thermal exposure the μ phase precipitating in the form of needles and granules experiences a complicated evolution. The needles, fiber- or sheet-shaped in three-dimension, align very regularly during exposure, either lying in three directions with an acute angle to one another or in two directions perpendicular to each other, the mechanisms of which are characterized in detail. In addition, it is concluded that an excessive precipitation of μ phase severely degrades the mechanical properties of the alloy, whereas its evolution behavior during thermal exposure is determined to have an insignificant influence on the properties.

Summary We have demonstrated spontaneous development of autoimmune cholangitis, similar to human primary biliary cirrhosis, in mice expressing a dominant negative form of the transforming growth factor-β receptor (dnTGF-βRII) restricted to T cells. The autoimmune cholangitis appears to be mediated by autoreactive CD8+ T lymphocytes that home to the portal tracts and biliary system. Because the liver pathology is primarily secondary to CD8+ T cells, we have determined herein whether administration of β-glucosylceramide (GC), a naturally occurring plant glycosphingolipid, alters the natural history of disease in this model. We chose GC because previous work has demonstrated its ability to alter CD8+ T cell responses and to down-regulate tissue inflammation. Accordingly, dnTGF-βRII mice were treated with either GC or control for a period of 18 weeks beginning at 6 weeks of age. Importantly, in mice that received GC, there was a significant decrease in the frequency and absolute number of autoreactive liver-infiltrating CD8+ T cells, accompanied by a significant decrease in activated CD44high CD8+ T cell populations. Further, there was a significant reduction in portal inflammation in GC-treated mice. Interestingly, there were no changes in anti-mitochondrial antibodies, CD4+ T cells, CD19+ B cells or natural killer (NK) T cell populations, indicating further that the beneficial effects of GC on liver inflammation were targeted specifically to liver-infiltrating CD8+ T cells. These data suggest that further work on GC in models of CD8+ T-mediated inflammation are needed and point to a new therapeutic venue for potentially treating and/or modulating autoimmune disease.

SummaryOur laboratory has suggested that loss of tolerance to pyruvate dehydrogenase (PDC-E2) leads to an anti-mitochondrial antibody response and autoimmune cholangitis, similar to human primary biliary cirrhosis (PBC). We have suggested that this loss of tolerance can be induced either via chemical xenobiotic immunization or exposure to select bacteria. Our work has also highlighted the importance of genetic susceptibility. Using the non-obese diabetic (NOD) congenic strain 1101 (hereafter referred to as NOD.1101 mice), which has chromosome 3 regions from B6 introgressed onto a NOD background, we exposed animals to 2-octynoic acid (2OA) coupled to bovine serum albumin (BSA). 2OA has been demonstrated previously by a quantitative structural activity relationship to react as well as or better than lipoic acid to anti-mitochondrial antibodies. We demonstrate herein that NOD.1101 mice immunized with 2OA-BSA, but not with BSA alone, develop high titre anti-mitochondrial antibodies and histological features, including portal infiltrates enriched in CD8+ cells and liver granulomas, similar to human PBC. We believe this model will allow the rigorous dissection of early immunogenetic cause of biliary damage.

Pharmacokinetics of difloxacin, a fluoroquinolone antibiotic, was determined in pigs and broilers after intravenous (i.v.), intramuscular (i.m.), or oral (p.o.) administration at a single dose of five (pigs) or 10 mg/kg (broilers). Plasma concentration profiles were analyzed by a compartmental pharmacokinetic method. Following i.v., i.m. and p.o. doses, the elimination half-lives (t(1/2beta)) were 17.14 +/- 4.14, 25.79 +/- 8.10, 16.67 +/- 4.04 (pigs) and 6.11 +/- 1.50, 5.64 +/- 0.74, 8.20 +/- 3.12 h (broilers), respectively. After single i.m. and p.o. administration, difloxacin was rapidly absorbed, with peak plasma concentrations (C(max)) of 1.77 +/- 0.66, 2.29 +/- 0.85 (pigs) and 2.51 +/- 0.36, 1.00 +/- 0.21 microg/mL (broilers) attained at t(max) of 1.29 +/- 0.26, 1.41 +/- 0.88 (pigs) and 0.86 +/- 0.4, 4.34 +/- 2.40 h (broilers), respectively. Bioavailabilities (F) were (95.3 +/- 28.9)% and (105.7 +/- 37.1)% (pigs) and (77.0 +/- 11.8)% and (54.2 +/- 12.6)% (broilers) after i.m. and p.o. doses, respectively. Apparent distribution volumes(V(d(area))) of 4.91 +/- 1.88 and 3.10 +/- 0.67 L/kg and total body clearances(Cl(B)) of 0.20 +/- 0.06 and 0.37 +/- 0.10 L/kg/h were determined in pigs and broilers, respectively. Areas under the curve (AUC), the half-lives of both absorption and distribution(t(1/2ka), t(1/2alpha)) were also determined. Based on the single-dose pharmacokinetic parameters determined, multiple dosage regimens were recommended as: a dosage of 5 mg/kg given intramuscularly every 24 h in pigs, or administered orally every 24 h at the dosage of 10 mg/kg in broilers, can maintain effective plasma concentrations with bacteria infections, in which MIC(90) are0.25 microg/mL and0.1 microg/mL respectively.

Summary Current models of adult haematopoiesis propose that haematopoietic stem cells (HSCs) differentiate into common lymphoid (CLP) and common myeloid (CMP) progenitors and establish an early separation between myeloid and lymphoid lineages. Nevertheless, the developmental potential of CMP-associated B cells suggests the existence of alternate pathways for B lymphopoesis. The aim of this study was to compare the developmental and functional properties of CMP- and CLP-derived B cells. While both populations matured through pro-B cell and transitional B cell intermediates in the bone marrow and spleen, respectively, following transfer into irradiated mice, mature CMP- and CLP-derived B cells exhibit distinct functional responses. Specifically, CMP-derived B cells did not respond to mitogenic stimulation to the same degree as their CLP-derived counterparts and secrete lower levels of IgM and the inflammatory cytokines such as interleukin (IL)-6 and IL-10. Together, these data suggest the existence of multiple pathways for generating functionally distinct B cells from bone marrow precursors.

Summary The role of the liver in the initiation and maintenance of tolerance is a critical immune function that involves multiple lineages of immune cells. Included within these populations are liver dendritic cells (DCs). Although there has been significant work on the phenotypic and functional roles of splenic and bone marrow dendritic cells, as well as their subsets, comparable studies in liver have often been difficult. To address this issue we have isolated, from C57BL/6 mice, relatively pure populations of DCs and compared phenotype and function to the data from spleen using flow cytometry, cell sorter assisted purification and culture, morphology by cytospin and May–Giemsa staining, cell cycle progression, antigen uptake, cytokine production and allo-activation potential. natural killer (NK)1·1–CD11c+ liver DC subsets (conventional DCs, T cell receptor (TcR)β–NK1·1–CD11c+B220– and plasmacytoid DCs, TcRβ–NK1·1–CD11c+B220+) efficiently endocytose dextran and produce significant levels of tumour necrosis factor (TNF)-α, interleukin (IL)-6 and IL-12 p40 in response to Toll-like receptor (TLR) ligands, with responses higher than splenic DCs. There is also a differential capability of hepatic DCs to respond to innate signals. Indeed, CD11c+ hepatic DCs have a greater capacity to respond to innate stimulation but are less capable of inducing CpG activated-allogeneic T cells. These data suggest that hepatic dendritic cells function as a critical bridge between innate and adaptive immunity and are capable of inducing stronger innate responses with a lower capacity for allo-stimulation than splenic dendritic cells. These properties of liver dendritic cells contribute to their unique role in the induction of tolerance.

Summary Several epidemiological studies have demonstrated that patients with primary biliary cirrhosis (PBC) have a higher incidence of urinary tract infections (UTI) and there is significant homology of the immunodominant mitochondrial autoantigen, the E2 component of the pyruvate dehydrogenase complex (PDC-E2), between mammals and bacteria. Previous work has demonstrated that non-obese diabetic (NOD).B6 Idd10/Idd18 infected with Novosphingobium aromaticivorans developed liver lesions similar to human PBC. It was postulated that the biliary disease was dependent upon the presence of the unique N. aro glycosphingolipids in activating natural killer T (NK T) cells. To address this issue, we infected NOD.B6 Idd10/Idd18 mice with either Escherichia coli, N. aro or use of a phosphate-buffered saline (PBS) vehicle control and serially followed animals for the appearance of liver pathology and anti-mitochondrial autoantibodies (AMA). Of striking importance, the biliary disease of E. coli-infected mice was more severe than N. Aro-infected mice and the titre of AMA was higher in E. coli-infected mice. Furthermore, the immunopathology did not correlate with the ability of bacterial extracts to produce antigen-dependent activation of NK T cells. Our data suggest that the unique glycosphingolipids of N. aro are not required for the development of autoimmune cholangitis. Importantly, the data highlight the clinical significance of E. coli infection in a genetically susceptible host, and we suggest that the appearance of autoimmune cholangitis is dependent upon molecular mimicry. These data highlight that breach of tolerance to PDC-E2 is probably the first event in the natural history of PBC in genetically susceptible hosts.

In order to understand the molecular mechanisms by which two types of phytohormones, brassinosteroids (BRs) and gibberellins (GAs), regulate plant growth and development, a cDNA microarray was made containing 4000 clones randomly selected from a rice cDNA library constructed from RNA isolated from seedlings treated with GA(3) and brassinolide (BL). The array was then probed to identify differences in expression using labelled cDNAs prepared from treated and control seedlings. Nine and 29 unique cDNA clones were up-regulated, while 32 and 42 unique cDNA clones were down-regulated by BL and GA(3), respectively. The predicted products of these BL- and GA-regulated genes fall into such functional categories as signal transduction, transcription, metabolism, cellular organization, and defense or anti-stress responses. Northern analysis of the selected BL- and GA(3)-regulated genes not only confirmed their BL or GA(3) dose dependent expression, but also revealed significant differences in the degree of expression in different organs and in response to other phytohormones. These results demonstrate that BR and GA influence growth and development by coordinately regulating the expression of specific groups of genes.

To study the relationship between duck hepatitis B virus (DHBV) infection and duck hepatocellular carcinoma (DHCC), histological examination and DHBV DNA hybridization were performed in 875 ducks from three flocks in Qidong County. Among them, 34 suffered from hepatoma, including 23 hepatocellular carcinoma, 8 cholangiocarcinoma and 3 hepatocellular-cholangiocarcinoma. Of the 34 ducks with hepatoma 27 were positive for DHBV DNA in the liver and/or serum. DHBV DNA was demonstrated in neoplastic nodules of 22 ducks. Southern blot analysis showed that 13 cases were of the integrated pattern of DHBV DNA in neoplastic nodules. The paratumor tissues of 14 ducks with massive tumor were analysed at the same time. Five cases showed integrated pattern, 4 cases free pattern and the other 4 cases both integration and free pattern of DHBV DNA. The hybridization pattern of DHBV DNA in tumor nodule was different from that in paratumor regions in 11 cases and identical in 3 cases. DHBV antigen was positive in 13 tumor nodules and 21 paratumor tissues in the 34 ducks with hepatic tumor by both victoria blue and orcein stain methods. Advanced liver diseases were found in 30 out of the 34 ducks with hepatoma, including 12 cirrhosis and 18 chronic active hepatitis. In southern blot analysis of 122 DHBV DNA positive Qidong ducks without hepatoma, only free pattern of DHBV was seen, while 44 control ducks from Changchun were negative for DHBV DNA. Neither hepatic tumor nor liver diseases were seen in the control ducks. The results suggest that hepatocellular carcinoma in ducks is similar to that in human HCC. They have a high frequency of viral DNA integrated into the host genome and a liver disease background.

The hybridization test for duck hepatitis B virus DNA was performed in 34 ducks with hepatoma from Qidong, Jiangsu province. Among the 34 hepatoma ducks, 18 were positive for DHBV DNA in the serum and 27 were positive in the tumor and/or liver tissue. Tissue sections were stained with Victoria blue nuclear fast red for detecting DHBV surface antigen. Victoria blue positive cells were found in 11 tumors and 15 paratumorous regions of 23 ducks with hepatocellular carcinoma. Although paratumorous regions were positively stained in 3 of 8 ducks with cholangiocarcinoma, all were negative within their tumors. All paratumorous regions and 2 tumor regions of 3 ducks with hepatocellular-cholangiocarcinoma were positive for Victoria blue. The results suggest that duck hepatocellular carcinoma be closely related to DHBV infection.

Duck hepatitis B virus (DHBV) DNA was detected in both multiple primary cancers and intrahepatic metastatic tumors in duck liver by Southern blot technique. Among 7 ducks with multiple primary tumors in the liver, the banding pattern of integrated DHBV DNA in different tumors was identical in 3 animals but different in 2. One duck showed a similar hybridization pattern in different tumor nodules and the remaining one was negative for DHBV DNA. Integrated DHBV DNA was also identified in one duck with metastatic tumor in the lung, in which the hybridization pattern was the same as that of the primary hepatocellular carcinoma. The presence of the same distinct hybridization bands in various tumor nodules indicated that they may arise from the same clone of neoplastically transformed cell. Different hybridization bands showed that tumors may arise from different clones. The results suggest that hybridization pattern of hepatic carcinoma with viral DNA probe could be used as a clone marker of transformed cells to distinguish multiple primary hepatic cancers from intrahepatic metastasis.

From 1979 to 1986, 121 patients with electrical burns were admitted to our, unit (6% of all the burned patients admitted during that period). Of these cases 64 were regarded as having deep burns (e.g. burns which injured muscles, tendons, nerves, bones and capsules of the joints etc.). Thirty-five flaps of various types of tissues (including one free muscle flap, two island fascial flaps, four island skin flaps, three arterial skin flaps, two musculocutaneous flaps and twenty-three random skin flaps) were used for the coverage of deep burns in twenty-seven patients. Thirty three flaps survived completely while two were partially lost; three flaps developed severe sub-flap infections but the tissue of the flaps survived totally. Three typical cases are reported in this paper. The advantages of early extensive exploration and thorough debridement of the deep electrical wounds are discussed. It also introduces the practical usage of various flaps for wound coverage in these typical cases. When flaps were needed for deep electrical burns, we chose local flaps first, and used axial pattern flaps, island flaps, etc. as much as possible; if only a random flap was available in the site, we elevated it with some deep fascia to enhance its ability to prevent infection. We considered the free tissue transfer and the transfer of fascial flaps to be important alternatives. Debridement followed by immediate wound coverage should be done as soon after resuscitation as possible. Generally the operations were performed within the third to fifth days after the electrical burn. This is the best period for active treatment in order to salvage the remaining functions of the burned extremities. Operations performed after six days must be done only under the careful control of anti-infective measures. From our experience we have found that muscle flaps and musculocutaneous flaps were the most suitable approaches for the coverage of deeply infected wounds especially when bones were involved. Careful attention was taken to excise the necrotic muscle tissues, especially those that were "sandwiched" between necrotic muscle bundles. This was done primarily by gross evaluation according to the judgement of the surgeon intraoperatively. The swollen and hardened regions of the skin of involved extremities generally corresponded to the areas of necrotic muscle tissue at about three days after the injuries or later. This was used as a reference to indicate where to place or extend the exploratory incisions. We frequently preserved denatured peripheral nerves and tendons, because these injured structures often later regain their functions partially or completely with a reliable coverage.(ABSTRACT TRUNCATED AT 400 WORDS)