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1.
Immunohistochemistry for E-cadherin (ECAD) has been used to distinguish canine cutaneous histiocytoma from other leukocytic neoplasms ("round cell tumors"). To determine the specificity of this test, 5 types of canine cutaneous round cell tumors were evaluated for immunohistochemical expression of ECAD. Tumors of all 5 types had variable cytoplasmic, plasma membrane, and/or paranuclear ECAD expression: All 13 cutaneous histiocytomas were ECAD+; all but 1 of 14 mast cell tumors expressed ECAD; 10 of 12 epitheliotropic lymphomas reacted with E-cadherin antibody; of 72 plasmacytomas, 54 were ECAD+; and 5 of 5 histiocytic sarcomas were positive. Conclusions based on these results include the following: First, immunoreactivity for ECAD is not limited to leukocytes of cutaneous histiocytoma; second, antibody to ECAD also labels neoplastic cells in most mast cell tumors, plasmacytomas, cutaneous histiocytic sarcomas, and epitheliotropic lymphomas; third, although most histiocytomas have membranous ECAD expression, the immunoreactivity varies among round cell tumors and is frequently concurrent in different cellular compartments; fourth, the distinctively paranuclear ECAD expression pattern in epitheliotropic lymphomas might distinguish them from other round cell tumors; and, fifth, ECAD should be used with other markers (eg, MUM1 for plasmacytomas, KIT for mast cell tumors, CD3 and CD79a for lymphomas) to distinguish among canine round cell tumors.  相似文献   

2.
Disorders of mast cells, particularly mast cell tumors (MCTs), are common in dogs. There now is evidence that many of these disorders exhibit breed predilections, suggesting an underlying heritable component. In comparison to humans and mice, little is known regarding the biology of canine mast cells. To facilitate the study of mast cell biology in other species, bone marrow-derived cultured mast cells (BMCMCs) often are used because these represent a ready source of large numbers of cells. We have developed a protocol to successfully generate canine BMCMCs from purified CD34(+) cells. After 5-7 weeks of culture with recombinant canine stem cell factor (rcSCF), greater than 90% of the cell population consisted of mast cells as evidenced by staining with Wright's-Giemsa, as well as production of chymase, tryptase, IL-8 and MCP-1. These cells expressed cell surface markers typical of mast cells including Kit, Fc epsilonRI, CD44, CD45 and CD18/CD11b. The canine BMCMCs were dependent on rcSCF for survival and proliferation, and migrated in response to rcSCF gradients. Cross-linking of cell surface-bound IgE induced the release of histamine and TNFalpha. Histamine release could also be stimulated by ConA, compound 48/80, and calcium ionophore. In summary, canine BMCMCs possess phenotypic and functional properties similar to mast cells found in vivo. These cells represent a novel, valuable resource for investigating normal canine mast cell biology as well as for identifying factors that lead to mast cell dysregulation in the dog.  相似文献   

3.
Among the intestinal tumors of hematopoietic cell origin, lymphoma is the most common in the dog. Herein, we characterized the clinical and pathologic features of 11 dogs (average age, 10.6 +/- 2.5 years) with T-cell lymphoma of the intestinal tract with eosinophil infiltrates. No sex predominance was apparent. All had localized tumor masses in the small intestine. Grossly, the intestinal wall was thickened, and the lumen of the affected intestine was usually narrowed. Microscopically, we observed transmural diffuse invasion of round to pleomorphic tumor cells. Tumor cells showed varying morphology, from scanty to abundant cytoplasm, and round to ovoid nuclei with scattered to dense chromatin. In seven of the dogs, tumor cells had infiltrated into the epithelium. All showed infiltration of eosinophils and all 11 tumors had a T-cell phenotype (CD3+, CD79-). Only one tumor stained positive for the mast cell marker c-kit and none was positive for mast cell tryptase. We did not observe ultrastructurally apparent granules in any of the tumor cells. These results suggest that, in dogs, T-cell lymphomas of intestinal origin resemble mast cell tumors of intestinal origin with respect to cell structure and eosinophil infiltration. Therefore, in the absence of epitheliotropism, it is difficult to confirm the differential diagnosis without immunostaining for mast cell and lymphocyte markers, including mast cell tryptase, c-kit, CD3, and CD79.  相似文献   

4.
Sixty-four canine cutaneous round cell tumors were divided into 25 mast cell tumors, 15 histiocytomas, nine cutaneous lymphosarcomas and 15 transmissible venereal tumors. The final diagnosis was made from cytologic, clinical and histologic findings. Cytologic features were significantly distinctive in mast cell tumor, transmissible venereal tumor, and most cases of histiocytoma and lymphosarcoma to allow a diagnostic opinion. This opinion was supported by subsequent histologic examination. In some instances cytology was considered essential in rendering a diagnostic opinion even though histology was available.  相似文献   

5.
Studies using the currently available malignant canine mast cell lines and bone marrow-derived cultured mast cells (BMCMCs) have provided an in-depth understanding of normal and neoplastic canine mast cell biology. However, many of the currently available malignant canine mast cell lines possess limitations, including loss of cell surface markers and inability to bind canine IgE. We have recently generated a novel mast cell line, CL1, from an 11-year-old spayed female Labrador retriever diagnosed with systemic mastocytosis and neoplastic effusion. The CL1 cells express KIT, Fc?RI, CD44, CD45, CD14, CD11a, CD11b and CD18 as well as chymase. Interestingly, these cells express wild-type KIT, with no evidence of autophosphorylation, but are able to proliferate independently without the addition of exogenous stem cell factor (SCF), KIT ligand. However, stimulation of CL1 cells with SCF induces KIT phosphorylation promoting cell proliferation. The CL1 cells retain functional properties of mast cells, degranulating in a dose-dependent manner in response to both IgE cross-linking and chemical stimulation. Lastly, cytogenetic evaluation revealed several recurrent tumor-associated chromosome copy number imbalances in the CL1 line. In summary, the CL1 cell line possesses phenotypic and functional properties similar to those found in canine BMCMCs, and will likely be a useful tool to study mast cell biology, factors regulating transformation of mast cells, cytogenetic abnormalities in mast cell tumors, and novel preclinical therapies.  相似文献   

6.
It has been suggested but not proven that hypersensitivity type I reactions are involved in the pathogenesis of canine inflammatory bowel disease (IBD). The main effector cells in type I hypersensitivity reactions are mast cells (MCs). Canine MCs, as human MCs, can be subdivided into three subtypes according to their content of mast cell-specific proteases: tryptase (MCT), chymase (MCC), or tryptase and chymase bearing MCs (MCTC). In this study, numbers and subsets of mast cells were investigated in biopsies from the gastrointestinal tract of dogs with histopathologically confirmed lymphocytic-plasmacytic enteritis (LPE) (n = 4), lymphocytic-plasmacytic colitis (LPC) (n = 1) and eosinophilic gastroenterocolitis (EGE) (n = 11). Paraffin sections of formalin-fixed samples from the stomach, small intestine (duodenum, jejunum, ileum) and colon were stained by using a metachromatic staining method (kresylecht-violet; KEV) and a combined enzyme histochemical and immunohistochemical technique for chymase and tryptase. Additionally, immunohistochemistry with antibodies against T cells (CD3), macrophages (myeloid/histiocyte antigen) and IgA, IgG and IgM bearing cells was conducted. Quantitative evaluation of mast cells and semiquantitative scoring of immunohistochemically stained cells were performed. Between the two histopathologically defined groups clear differences concerning mast cell numbers were detected. In most affected intestinal tissue locations of dogs with LPE/LPC a decrease in metachromatically (kresylecht-violet) stained granule-containing MCs and immunohistochemically stained MCT,C,TC was found. This reduction could be due to mast cell degranulation, a T helper cell 1 dominated reaction pattern or a “thinning out” due to increasing T cells, IgA and IgG bearing cells. Dogs with EGE displayed higher variability in mast cell numbers but most of the affected large and small intestinal locations had increased numbers of MCs. In these cases, T cells, IgA bearing cells and macrophages also increased. Increased numbers of MCs and eosinophils seen in the intestinal mucosa of dogs with EGE could indicate the presence of a type I hypersensitivity reaction (T helper cell 2 pattern) in response to dietary antigens. Changes in cell numbers occurred also in unaffected locations of dogs with LPE/LPC and EGE which showed reduced MCT,C,TC, increased KEV positive cells and partially increased leucocytes and macrophages.  相似文献   

7.
The objective of this study was to evaluate by immunohistochemical means the nuclear expression of p27 and p21 proteins in cutaneous mast cell tumors and histiocytomas of dogs. In mast cell tumors, nine of the 13 grade I tumors, 13 of the 19 grade II tumors, and 10 of the 15 grade III tumors showed no detectable or mild p27 immunoreactivity. In contrast, one of the 13 grade I tumors, 12 of the 19 grade II tumors, and 11 of the 15 grade III tumors showed moderate or marked p21 immunoreactivity. Nineteen of the 28 histiocytomas showed no detectable or mild p27 immunoreactivity, and 24 cases showed moderate or marked p21 immunoreactivity. These findings indicate that a loss or absence of p27 expression is an early pathogenic event in mast cell and histiocyte tumorigenesis and that p21 expression may be a marker of mast cell tumor progression and histiocytoma cell proliferation.  相似文献   

8.
Gain-of-function mutations in the proto-oncogene c-kit have been considered the molecular mechanism of neoplastic proliferation of mast cells. However, the importance of c-kit gene mutations is not well evaluated in canine mast cell tumors (MCTs). In the present study, we established and characterized a mast cell line, HRMC, derived from a dog with MCT. We also examined c-kit mutations in HRMC cells and assessed an inhibitory effect of a tyrosine kinase inhibitor, STI571, on HRMC cells. HRMC cells had cytoplasmic metachromatic granules, chymase and tryptase, and expressed both KIT and FcepsilonRI on the cell surface. HRMC cells contained histamine and released beta-hexosaminidase through FcepsilonRI cross-linking and calcium ionophore stimulation. Nucleotide sequence analysis demonstrated no mutations in an open reading frame of c-kit cDNA and genomic DNA of the juxtamembrane domain of c-kit in HRMC cells. STI571 did not show any inhibitory effects on the proliferation of HRMC cells. These findings clearly demonstrated the existence of c-kit mutations-independent neoplastic canine mast cell proliferation. The growth factor-independent mast cell line established in this study might be valuable to explore novel mechanisms of c-kit mutations-independent neoplastic proliferation of mast cells in dogs.  相似文献   

9.
Mast cell tumors (MCTs) of gastrointestinal origin that had been surgically removed from 39 dogs were examined to evaluate their pathologic features. Miniature breeds, especially Maltese, were most frequently affected. The average age of affected dogs was 9.7 +/- 2.6 years. No sex difference was apparent. The most frequently affected sites were in the upper digestive tract, and the prognosis was very poor. Grossly, the gastrointestinal wall was prominently thickened, and the lumen of the affected gut was usually narrowed. Microscopically, there was diffuse transmural invasion of round to pleomorphic tumor cells. Tumor cells had moderate to abundant cytoplasm, round to ovoid nuclei with scattered chromatin, and mitotic figures. Fibrous stroma was observed in about half of the tumors. There was variable infiltration of eosinophils. In all tumors, cytoplasmic granules showed weak metachromasia, but the number of granules was very small. Immunohistochemical staining for c-kit and mast cell tryptase was positive in 77% and 62% of tumors, respectively. All tumors were positive for at least two of these markers. Immunohistochemical staining for p53 was positive in 13% of the tumors. Reactivity for staining markers and p53 was unrelated to cell pleomorphism, vessel invasion, or survival time. Gastrointestinal MCTs have histologic and immunohistochemical features completely different from those of other primary or metastatic gastrointestinal tumors. The combination of immunostaining for mast cell tryptase and c-kit and histochemical staining for metachromasia appears to be a powerful tool for the diagnosis of gastrointestinal MCTs.  相似文献   

10.
Sixty-five canine skin neoplasms studied using immunocytochemistry, included 22 histiocytomas, 18 amelanotic melanomas, 14 cutaneous lymphosarcomas, six mast cell tumors, and five transmissible venereal tumors. Formalin-fixed, paraffin-embedded sections were stained using the avidin-biotin-peroxidase complex (ABC) immunoperoxidase technique for reactivity with S-100 protein, kappa and lambda immunoglobulin light chains, alpha-1-antitrypsin, alpha-1-antichymotrypsin, leukocyte common antigen (LCA), neuron-specific enolase, keratin, cytokeratin, muramidase, and vimentin. Detection of S-100, kappa and lambda light chains, neuron-specific enolase, and vimentin were most useful for screening these neoplasms. None of the markers examined was consistent in staining histiocytomas. While reactivity of S-100 (ten cases) and neuron-specific enolase (ten cases) was detected in some amelanotic melanomas, lambda light chain immunoglobulin (eight cases) was relatively consistent in cutaneous lymphomas. Mast cell neoplasms reacted with avidin and, therefore, were positive, even on negative control sections. Vimentin reacted strongly on all amelanotic melanomas and transmissible venereal tumors examined. These antibodies are helpful adjuncts in the differential diagnosis of canine skin tumors.  相似文献   

11.
Disorders involving mast cells are extremely common in dogs, ranging from allergic diseases to neoplastic transformation resulting in malignant mast cell tumors. Relatively little is known regarding the basic biologic properties of normal canine mast cells, largely due to the difficulty in reliably purifying large numbers from canine skin. In vitro generated bone marrow derived cultured mast cells (BMCMCs) are routinely used in both human and murine studies as a ready source of material for in vitro and in vivo studies. We previously developed a technique to generate canine BMCMCs from bone marrow derived CD34+ cells and demonstrated that these cells exhibit the phenotypic properties characteristic of mast cells and release histamine in response to IgE cross-linking. The purpose of the following study was to characterize the functional properties of these canine BMCMCs and contrast these with the functional properties of murine BMCMCs. Our work demonstrates that both IL-4 and IL-10 promote canine BMCMC proliferation, possibly through upregulation of Kit expression, while TGFbeta inhibits proliferation. The canine BMCMCs produce a variety of cytokines and chemokines in response to IgE cross-linking and chemical stimulation including IL-3, IL-4, IL-13, GM-CSF, RANTES, and MIP1alpha. Interestingly, the canine BMCMCs released significantly larger amounts of MCP-1 and tryptase and significantly smaller amounts of IL-6 following chemical stimulation and IgE cross-linking when compared to murine BMCMCs. Lastly, the canine BMCMCs produced larger amounts of active MMP9 than their murine counterparts. In summary, canine BMCMCs exhibit unique functional properties that distinguish them from murine BMCMCs and provide insight into the contribution of these cells to mast cell disorders in the dog.  相似文献   

12.
Vascular endothelial growth factor (VEGF) and prostaglandin E2 (PGE-2) appear to play a critical role in tumour neovascularization. In this study, we have investigated the expression of VEGF and PGE-2 in 53 canine cutaneous mast cell tumours (MCTs). Immunohistochemistry of tissue sections revealed that VEGF and PGE-2 were expressed in all mast cell tumours studied. When the expression patterns of VEGF and PGE-2 were compared with tumour grade according to Patnaik criteria, the only significant correlation observed was between PGE-2 staining intensity and tumour pathological grade, with grade II and III tumours having higher PGE-2 staining, both in intensity and percentage of cells stained, than grade I tumours ( P < 0.05).  相似文献   

13.
A new cell line (CoMS) was established from a 3-year-old male mongrel dog with mast cell tumor of the oral mucosa. CoMS cells grow in suspension with a doubling time of 27.0 +/- 0.7 hr. The cytoplasmic granules were formalin-sensitive, showed diverse appearances in their ultrastructural findings and contained heparin proteoglycan and neutral protease chymase. Calcium ionophore A23187, substance P and concanavalin A caused significant histamine release from CoMS cells, while compound 48/80 failed to release histamine. This cell line will make an available source for studies on canine mast cell tumors.  相似文献   

14.
Presence of matrix metalloproteinases has been associated with tumor invasion and metastasis in human neoplasia. The presence of matrix metalloproteinase 2 and matrix metalloproteinase 9 was determined in canine mast cell tumor tissue and normal stromal tissue from 24 dogs with spontaneously occurring cutaneous mast cell tumors. Seventeen of the mast cell tumors were of histologic grade 2, and 7 were of histologic grade 3. Gelatin zymography and computer assisted densitometry image analysis were used to quantify matrix metalloproteinase concentration. Bands from canine tissues migrated in the same location as human proenzyme and active enzyme matrix metalloproteinase 2 and matrix metalloproteinase 9 standards. A semiquantitative value for each patient sample was obtained by comparing the optical assessment density of each unknown band to the optical density of the human standard. The presence of matrix metalloproteinase 2 and matrix metalloproteinase 9 in histologic grade 2 mast cell tumors and histologic grade 3 mast cell tumors was compared, as was presence of matrix metalloproteinases in tumor and stromal tissue. There was dramatically more proenzyme matrix metalloproteinase 9 activity in histologic grade 3 mast cell tumors when compared to grade 2 tumors (P = .03). There was also dramatically more active enzyme matrix metalloproteinase 2 and active enzyme matrix metalloproteinase 9 activity in tumor tissue compared to stromal tissue (P = .02, P < .0001). This study demonstrates that the proenzyme and active enzyme forms of matrix metalloproteinase 2 and matrix metalloproteinase 9 are present in canine mast cell tumors. This appears to be related to the degree of histologic malignancy, although histologic grade 1 tumors were not evaluated.  相似文献   

15.
Thirty-one canine cutaneous masses, diagnosed as mast cell tumors (MCT) by histopathologic analysis, were used to evaluate the immunohistochemical pattern of expression of KIT protein (CD117), a type III tyrosine kinase protein involved in mast cell growth and differentiation. Lesions were graded as I (well differentiated), II (intermediate differentiation), or III (poorly differentiated) according to the following morphologic features: invasiveness, cellularity and cellular morphology, mitotic index, and stromal reaction. Immunohistochemical KIT expression was compared with histologic grade and some histomorphologic features (cell differentiation and nuclear grade) evaluated separately. A possible predictive role of biologic behavior in MCTs for KIT expression was also investigated. Immunohistochemical analysis revealed three different patterns of KIT expression: a cytoplasmic diffuse pattern, a membranous pattern with immunostaining located on the cell surface, and a cytoplasmic perinuclear pattern, where KIT expression was detected in the cytoplasm of the neoplastic mast cells, close to the nucleus. Statistical analysis showed a close relationship between different KIT immunohistochemical patterns and histologic grade (P < 0.00000), cell differentiation (P < 0.00000), and nuclear grade (P < 0.0024). According to Kaplan-Meier-estimated survival curves compared by survival analysis, KIT expression was significantly associated with survival time (P = 0.037) but not cancer-free interval (P = 0.50). Similar to other well-known histomorphological features, KIT expression is a useful parameter of malignancy in cutaneous MCTs. KIT expression also predicted the biological behavior of the tumors in this study.  相似文献   

16.
Cytologic sampling of the ultrasonographically normal spleen and liver is not implemented routinely in the clinical staging of canine cutaneous mast cell tumors and normal ultrasound findings are often accepted as sufficient evidence for ruling out splenic or liver metastasis. Our objective was to define the specificity and sensitivity of ultrasound findings for diagnosis of mast cell infiltration when verified with cytologic evaluation, and to define the prognostic role of cytologic evaluation of liver and splenic aspirates. Dogs with a diagnosis of clinically aggressive grade II, or grade III mast cell tumor treated with a combination vinblastine/CCNU chemotherapy protocol, were selected retrospectively based on availability of cytologic evaluation of spleen plus or minus liver for staging. Out of 19 dogs, 10 dogs had a grade II tumor and nine a grade III tumor. Seven dogs had mast cell infiltration of the spleen, liver, or both. The sensitivity of ultrasound for detecting mast cell infiltration was 43% for the spleen and 0% for the liver. Dogs with positive cytologic evidence of mast cell infiltration to spleen, liver, or both had significantly shorter survival (100 vs. 291 days) than dogs without evidence of mast cell infiltration (P<0.0001). Routine splenic aspiration should be performed regardless of ultrasonographic appearance in dogs with a clinically aggressive mast cell tumor.  相似文献   

17.
Flat-Coated Retrievers seem to be at increased risk of developing soft-tissue sarcomas, and undifferentiated round cell or spindle cell sarcomas account for approximately 59% of sarcomas in the breed. In an attempt to classify these tumors further, formalin-fixed, paraffin-embedded sections from 14 undifferentiated sarcomas from Flat-Coated Retrievers were reviewed and examined with a panel of histologic and immunohistochemical stains. The panel included vimentin, desmin, Myo D1, smooth muscle actin, cytokeratin, S100, von Willebrand factor (factor VIII), Mac 387, CD3, major histocompatibility complex (MHC) class II, and CD79a. The majority of the sarcomas showed greater than 70% staining for MHC class II. We conclude that these undifferentiated sarcomas in Flat-Coated Retrievers belong to a spectrum of tumors with varying proportions of characteristic cell types and morphologic features, some of which fit the diagnostic criteria for malignant fibrous histiocytoma. Many of these sarcomas seem to have a significant myofibroblast component and a mild or moderate T cell infiltrate but the precise cell lineage is still uncertain.  相似文献   

18.
OBJECTIVE: To compare the expression of major histocompatibility complex (MHC) class II antigen in the corneas of normal dogs and dogs affected with chronic superficial keratitis (CSK). METHODS: MHC class II expression was determined in frozen sections of normal canine cornea and cornea from lesions of CSK by immunohistochemistry using a monoclonal antibody directed against the canine MHC class II molecule. Langerhans cell phenotype was determined morphologically and by histochemical determination of ATPase activity. To determine the influence of gamma interferon on expression of MHC class II molecules by corneal cells, corneal explants were cultured with the cytokine and MHC class II expression determined as above. RESULTS: Numerous MHC class II-expressing cells were demonstrated within the stroma and epithelium of the normal corneal limbus and conjunctival epithelium while very little MHC class II expression was detected in the central region of normal canine cornea. In limbal and conjunctival epithelium, cells expressing MHC class II antigen showed ATPase activity, suggesting that they were Langerhans cells. Corneas from dogs with CSK showed MHC class II expression associated with stromal cells, some of which exhibited a dendritic morphology while most were lymphocytic. Corneal epithelial cells within the lesion also aberrantly expressed MHC class II. Corneal explants expressed MHC class II to varying degrees after differing periods of incubation with the cytokine gamma interferon. CONCLUSIONS: While the normal central cornea has little MHC class II expression, aberrant expression occurs in CSK, associated with secretion of gamma interferon by infiltrating CD4-expressing lymphocytes. Although this change is likely to be a secondary feature of the CSK lesion, increased MHC class II expression may play a part in perpetuating the corneal inflammation seen in the disease.  相似文献   

19.
Fifty-four canine cutaneous mast cell tumors were evaluated immunohistochemically for the expression of P-glycoprotein (PGP) and multidrug-resistance-associated protein (MRP). All tumors examined were graded according to the histological malignancy. ranging from grade I to III. The expression of PGP was confirmed in 15% (8/54) of whole, 33% (5/15) of grade I, 10% (3/31) of grade II, and 0% (0/8) of grade III tumors. The expression of MRP was found in 18% (10/54) of whole, 26% (4/15) of grade I, 19% (6/31) of grade II, and 0% (0/8) of grade III tumors. The cases positive to at least one of these 2 multidrug markers were 26%, 47%, 23% and 0% of whole and grade I to III tumors, respectively. These results indicate that at least 26% of canine cutaneous mast cell tumors express PGP and/or MRP and that these tumors may be resistant to several anti-cancer drugs.  相似文献   

20.
MHC class II proteins present fragments of extra cellular antigen to stimulate CD4(+) T lymphocytes. Aim of this study was the detection of MHC class II antigens on different cutaneous cells in canine demodicosis. Histopathological and immunohistochemical examination of skin biopsies from 44 dogs with demodicosis is reported. The control group consisted of skin biopsies taken from 10 necropsied dogs without obvious skin lesions. The immunohistological assessment of the MHC class II expression revealed MHC class II proteins on different cell types of infiltrating inflammatory cells, i.e. APCs (antigen-presenting cells), macrophages, T lymphocytes and B lymphocytes. The plasma cells, however, only showed expression in 32 (73%) of 44 cases. Generally it was noticeable that most plasma cells but never all of them expressed MHC class II. Neutrophils, mast cells and eosinophils were MHC class II negative. Furthermore, in 39 biopsies (89%) from dogs with demodicosis MHC class II positive follicular keratinocytes were found. The control group did not show MHC class II expression on epithelial cells. Concerning the endothelial cells, a total of 25 biopsies (57%) showed MHC class II expression in which different vascular plexuses were affected by staining. This examination shows that MHC class II expression in the skin of dogs suffering form demodicosis is elevated. Especially the MHC class II expression by follicular keratinocytes seems to be conspicuous. We hypothesize that this is in association with the development and the maintenance of follicular inflammation.  相似文献   

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