Modulation of CD163 receptor expression and replication of porcine reproductive and respiratory syndrome virus in porcine macrophages

Porcine reproductive and respiratory syndrome virus (PRRSV) has a specific cell tropism for differentiated macrophages, such as porcine alveolar macrophages (PAMs). We analyzed the expression of CD163 on PAMs and macrophages derived from CD14 positive blood monocytes (MDMs), in correlation with PRRSV replication. By flow cytometry analysis, we showed that the levels of CD163 expression correlated well with the overall level of PRRSV replication. We further examined the effects of modulators of macrophage function, including 12-O-tetradecanoylphorbol-13-acetate (TPA), lipopolysaccharide (LPS), and interleukin (IL)-10 on the expression of CD163 and PRRSV replication. Pre-treatment of PAMs or MDMs with TPA or LPS resulted in decreased expression of CD163 and reduction in PRRSV replication. On the contrary, the incubation of CD14 positive monocytes with IL-10 during differentiation into MDMs resulted in up-regulated expression of CD163 with a corresponding increase in PRRSV infection. These data indicate that the expression of CD163 on macrophages in different microenvironments, in vivo, may determine the replication efficiency and subsequent pathogenecity of PRRSV.     

Immunohistochemical expression of CD117 and vascular endothelial growth factor in retinoblastoma: possible targets of new therapies

CD117 (C-kit) is thought to play an important role in tumourigenesis. There are limited data in the literature concerning C-kit expression in retinoblastoma. To date, no immunohistochemical studies have been performed to assess the possible association of C-kit with vascular endothelial growth factor (VEGF) in retinoblastoma. This study was designed to investigate C-kit and VEGF immunoexpression in retinoblastoma, their relationship with prognostic parameters as well as the correlation between them. A prospective immunohistochemical study was conducted on 56 retinoblastoma cases. Patients who had received preoperative chemotherapy were excluded. Positive C-kit and VEGF immunoreactivity was observed in 48.2% and 76.8% of retinoblastoma cases respectively. No C-kit immunostaining was seen in the adjacent uninvolved retina. However, VEGF expression was detected within its vasculature. Retinoblastomas with combined pattern of tumour growth revealed a highly significant positive C-kit expression (P = 0.002) compared to cases with endophytic or exophytic growths. Also, positive C-kit expression was statistically higher in cases with optic nerve invasion (P = 0.001) and choroidal invasion (P ≤ 0.01) compared to negative cases. A highly significant positive VEGF expression was detected in cases with optic nerve invasion (P = 0.013) compared to negative cases. Moreover, a highly significant positive correlation was detected between C-kit and VEGF expression (P = 0.006). C-kit is a feature of more aggressive retinoblastomas, with increased expression in tumours spreading beyond the retina. Moreover, VEGF is vastly expressed in retinoblastoma and is associated with optic nerve invasion. Both C-kit and VEGF may represent potential therapeutic targets for retinoblastomas.     

Exogenous surfactant protects against endotoxin induced acute respiratory distress syndrome in rodents via vascular endothelial growth factor

Vascular endothelial growth factor (VEGF) is a potent angiogenic factor which is abundantly expressed in the normal lung and is released by numerous cell types. Using a bacteria-induced lung injury model and surfactant therapy in rats, VEGF expression in lung was investigated.Sprague Dawley male rats were divided into four groups: buffer controls; rats challenged with LPS (055:B5 E. coli); challenged with LPS and treated with porcine surfactant (P-SF); and challenged with LPS and treated with synthetic surfactant (S-SF). The expressions of VEGF, PCNA, and BrdU were studied.VEGF protein expression was decreased in comparison to the control rats, as seen by both Western immunoblot and immunohistochemistry. Protein expression of PCNA and proliferation index as determined by both PCNA and BrdU immunostaining were also seen to be decreased in the LPS-treated animals, and with the surfactant treatment the expression was increased.The downregulation of VEGF in the alveolar space may reflect the recovery from acute lung injury, which leads to the limited endothelial permeability, and may participate in the decrease in capillary number, as observed during acute respiratory distress syndrome with potentially significant clinical consequences.     

Effects of human placental lactogen on the expression of CD163 and CD14 on human monocytes in culture

The effect of human placental lactogen (hPL), a member of the somatomammotrophin family, on the regulation of the scavenger receptor molecules CD14 and CD163 on human monocytes cultured for 48h was investigated. Cells were cultured in the presence or absence of the hormone and also in the presence or absence of IFN-gamma and dexamethasone. Monocytes cultured in the presence of hPL showed a significant increase in the expression of CD14 in both males and females compared to background. When IFN-gamma and dexamethasone were added to the cultures, CD14 expression was decreased and was not rescued by the presence of hPL. hPL alone had no effect on the expression of CD163 on cultured monocytes from either gender, although cells cultured in the presence of IFN-gamma and dexamethasone showed a profound increase in their expression of CD163. This expression was augmented further by the presence of hPL in the cultures over a 48-h period. These results support the hypothesis of a potential role of this hormone in the regulation of the innate immune response.     

PK-15 cells transfected with porcine CD163 by PiggyBac transposon system are susceptible to porcine reproductive and respiratory syndrome virus

The PiggyBac (PB) transposon system is a non-viral DNA-transfer system in which a transposase directs integration of a PB transposon into a TTAA site in the genome. Transgenic expression of porcine CD163 is necessary and sufficient to confer non-permissive cells susceptible to infection with porcine reproductive and respiratory syndrome virus (PRRSV). Such permissive cells can be used as a tool for PRRSV cellular receptor and other studies. One of the problems in studying PRRSV is the lack of porcine cell lines. In this study, efficient transfection and expression of porcine CD163 in PK-15 cells by PB transposition was demonstrated. The stable PK-15^CD163 cell line was used in PRRSV infection assays. The data indicated that the average PB transgene copy number per genome was approximately 10. In line with previous literature the integration of PB into the genome had a bias toward the TTAA chromosomal site. The PK-15^CD163 cell line was susceptible to infection by different PRRSV strains and the virus grew to similar titers compared to the Marc-145 cell line. This simplification of PK-15^CD163 cell line production will provide a valuable tool to facilitate PRRSV cellular receptor studies and to accelerate existing vectors for PK-15 cell-based gene transfer and expression.     

Elucidating the molecular physiopathology of acute respiratory distress syndrome in severe acute respiratory syndrome patients

Acute respiratory distress syndrome (ARDS) is a severe form of acute lung injury. It is a response to various diseases of variable etiology, including SARS-CoV infection. To date, a comprehensive study of the genomic physiopathology of ARDS (and SARS) is lacking, primarily due to the difficulty of finding suitable materials to study the disease process at a tissue level (instead of blood, sputa or swaps). Hereby we attempt to provide such study by analyzing autopsy lung samples from patient who died of SARS and showed different degrees of severity of the pulmonary involvement. We performed real-time quantitative PCR analysis of 107 genes with functional roles in inflammation, coagulation, fibrosis and apoptosis; some key genes were confirmed at a protein expression level by immunohistochemistry and correlated to the degree of morphological severity present in the individual samples analyzed. Significant expression levels were identified for ANPEP (a receptor for CoV), as well as inhibition of the STAT1 pathway, IFNs production and CXCL10 (a T-cell recruiter). Other genes unassociated to date with ARDS/SARS include C1Qb, C5R1, CASP3, CASP9, CD14, CD68, FGF7, HLA-DRA, IGF1, IRF3, MALAT-1, MSR1, NFIL3, SLPI, USP33, CLC, GBP1 and TAC1. As a result, we proposed to therapeutically target some of these genes with compounds such as ANPEP inhibitors, SLPI and dexamethasone. Ultimately, this study may serve as a model for future, tissue-based analyses of fibroinflammatory conditions affecting the lung.     

CD163-positive cancer cells are a predictor of a worse clinical course in lung adenocarcinoma

CD163 is one of the scavenger receptors expressed on macrophages. However, several immunohistochemical studies have demonstrated that CD163 is also detected on cancer cells, and is associated with a poor prognosis. In the present study, we detected CD163 staining on cancer cells in lung adenocarcinoma and squamous cell carcinoma (SCC), and investigated the relationship between CD163 on cancer cells and the clinical prognosis. CD163 staining was seen in 128 of 342 adenocarcinoma cases and 35 of 103 SCC cases. Among the lung adenocarcinoma cases, the progression-free survival and overall survival were significantly shorter in the CD163 high group than the CD163 low group. A similar trend was observed among the SCC cases, but the difference was not statistically significant. Additionally, a higher number of macrophages was detected in areas with CD163-positive cancer cells when compared to areas with CD163-negative cancer cells. In summary, we found that CD163-positive cancer cells are a predictor of a worse clinical course in lung adenocarcinoma and SCC.     

Angiogenic and inflammatory markers in acute respiratory distress syndrome and renal injury associated to A/H1N1 virus infection

Acute kidney injury (AKI) is often associated to acute respiratory distress syndrome (ARDS) due to influenza A/H1N1 virus infection. The profile of angiogenic and inflammatory factors in ARDS patients may be relevant for AKI. We analyzed the serum levels of several angiogenic factors, cytokines, and chemokines in 32 patients with A/H1N1 virus infection (17 with ARDS/AKI and 15 ARDS patients who did not developed AKI) and in 18 healthy controls. Significantly higher levels of VEGF, MCP-1, IL-6, IL-8 and IP-10 in ARDS/AKI patients were detected. Adjusting by confusing variables, levels of MCP-1 ≥ 150 pg/mL (OR = 12.0, p = 0.04) and VEGF ≥ 225 pg/mL (OR = 6.4, p = 0.03) were associated with the development of AKI in ARDS patients. Higher levels of MCP-1 and IP-10 were significantly associated with a higher risk of death in patients with ARDS (hazard ratio (HR) = 10.0, p = 0.02; HR = 25.5, p = 0.03, respectively) even taking into account AKI. Patients with influenza A/H1N1 infection and ARDS/AKI have an over-production of MCP-1, VEGF and IP-10 possibly contributing to kidney injury and are associated to a higher risk of death.     

CD163 favors Mycobacterium leprae survival and persistence by promoting anti‐inflammatory pathways in lepromatous macrophages

Lepromatous macrophages possess a regulatory phenotype that contributes to the immunosuppression observed in leprosy. CD163, a scavenger receptor that recognizes hemoglobin–haptoglobin complexes, is expressed at higher levels in lepromatous cells, although its functional role in leprosy is not yet established. We herein demonstrate that human lepromatous lesions are microenvironments rich in IDO+CD163+. Cells isolated from these lesions were CD68+IDO+CD163+ while higher levels of sCD163 in lepromatous sera positively correlated with IL‐10 levels and IDO activity. Different Myco‐bacterium leprae (ML) concentrations in healthy monocytes likewise revealed a positive correlation between increased concentrations of the mycobacteria and IDO, CD209, and CD163 expression. The regulatory phenotype in ML‐stimulated monocytes was accompanied by increased TNF, IL‐10, and TGF‐β levels whereas IL‐10 blockade reduced ML‐induced CD163 expression. The CD163 blockade reduced ML uptake in human monocytes. ML uptake was higher in HEK293 cells transfected with the cDNA for CD163 than in untransfected cells. Simultaneously, increased CD163 expression in lepromatous cells seemed to be dependent on ML uptake, and contributed to augmented iron storage in lepromatous macrophages. Altogether, these results suggest that ML‐induced CD163 expression modulates the host cell phenotype to create a favorable environment for myco‐bacterial entry and survival.     

Elevated levels of soluble CD163 in sera and fluids from rheumatoid arthritis patients and inhibition of the shedding of CD163 by TIMP-3

The aim of the present study was to evaluate levels of soluble CD 163 in sera and fluids from rheumatoid arthritis (RA) patients and elucidate the mechanism that regulates the shedding of CD163. Levels of soluble CD163 in sera and fluids from RA patients were examined by a sandwich enzyme immunoassay and Western blotting. To determine the effects of tissue inhibitors of metalloproteinase (TIMPs) on the shedding of CD163 from monocytes/macrophages, levels of soluble CD163 in cultures of monocytes/macrophages and the expression of CD163 on monocytes/macrophages in the presence or absence of TIMPs were examined by a sandwich enzyme immunoassay and flow cytometry, respectively. The clinical marker that was most associated with serum levels of soluble CD163 was levels of CRP. TIMP-3, but not TIMP-1 or TIMP-2, inhibited the shedding of CD163 from monocytes/macrophages. It was shown that serum levels of soluble CD163 are a sensitive and reliable marker to monitor activated macrophages in synovitis from RA patients and the results imply that the responsible proteinase for the shedding of CD163 is not a member of the matrix metalloproteinases, but is likely to be a member of ADAMs.     

SARS肺组织中CD34、VEGF、ICAM-1的表达及其意义

目的 通过对严重急性呼吸综合征(SARS)肺组织中CD34、血管内皮生长因子(VEGF)和细胞问黏附分子(ICAM-1)表达水平的检测对其发病机制进行探讨.方法 将7例死亡SARS病例及同期6例具有急性肺损伤性临床表现的非SARS死亡病例肺标本对照进行HE染色及CD34、VEGF、ICAM-1免疫组化检查,并对免疫组化染色结果进行图像分析.结果 CD34染色结果显示SARS组肺毛细血管内皮细胞着色浓重胞膜完整、排列紊乱但细胞连续性好,平均光密度值(MOD)为0.284 3±0.103 3;非SARS组肺组织CD34染色浅淡、不连续,MOD值为0.129 7±0.007 3,两组结果差异有显著性(P=0.007 4);不合并细菌或真菌感染SARS标本的MOD值为0.342 8±0.031 6,与非SARS组结果差异有显著性(P＜0.000 1).SARS组VEGF阳性染色MOD为0.125 3±0.042,与非SARS组0.114 3±0.025 2相比,结果差异无统计学意义(P=0.5888).两组ICAM-1阳性染色MOD值差异无统计学意义;但不合并感染的SARS标本ICAM-1 MOD值为0.255 7±0.045 2,与非SARS组0.182±0.017相比,结果差异有统计学意义(P=0.006).结论 SARS肺脏中内皮细胞的损伤程度较之其他原因引起的肺组织损伤轻,ICAM.1和VEGF表达也存在不同,这些差异可能反映了SARS肺损伤的特殊性,对解释SARS的临床和病理生理过程具有一定意义.     

结直肠癌CD44V6、E-cadherin和VEGF的表达及其意义

目的：探讨结直肠癌CD44V6、E-cadherin和VEGF的表达及其临床意义。方法：应用免疫组化S－P法检测46例结直肠癌CD44V6、E-cadherin和VEGF的表达,并分析其与结直肠癌临床病理特征的关系。同时检测了23例淋巴结转移灶CD44V6、E-cadherin和VEGF的表达。结果：结直肠癌组织中CD44V6、E-cadherin和VEGF阳性表达率分别为50．0％、56．5％、和50．0％;原发灶CCD4V6和VEGF阳性表达率低于淋巴结转移灶（60．9％和56．％）,而原发灶E-cadherin阳性表达率则高于淋巴结转移灶（43．5％）。结直肠癌CDV46和VEGF阳性表达与淋巴结转移,局部浸润深度和Dukes分期等有关。E-cadherin阳性表达与结直肠癌临床病理特征无关。结论：CD44V6和VEGF是结直肠癌发生与发展重要的促进因子,其过度表达与结直肠癌浸润转移,病理分期有关,可作为预测结直肠癌预后的生物学指标之一。     

Enhancement of CD4, CD5, CD8, and Bcl-2 Immunohistochemical Staining with Biotinyl-Tyramide

Abstract

Sections of formalin fixed, paraffin embedded tonsil were reacted with antibody for CD4, 5, and 8, and Bcl-2 followed by biotinylated secondary antibodies and ABC development. For each stain, parallel sections were either enhanced with biotinyl tyramide or not. Although there was a small increase in nonspecific background, the enhancement of the specific stain enabled a clearer identification of positive cells and in some cases increased the number of positive cells seen. (The J Histotechnol 21:237–239, 1998)     

血清IMA、MCP-1、CD163表达水平与急性冠脉综合征患者病情严重程度的关系研究

目的研究血清IMA、MCP-1、CD163表达水平与急性冠脉综合征患者病情严重程度的关系。方法本研究的研究对象以2017年1月至2018年1月在我院诊治的100例ACS患者为观察组,同期100例非缺血性胸痛患者为对照组,分别对两组患者的血清IMA、MCP-1、CD163表达水平进行对比,对观察组患者的以上指标进行多因素相关回归分析。结果观察组患者的IMA、MCP-1、CD163水平与Gensini评分、血浆NT-proBNP水平明显高于对照组,差异具有统计学意义(P <0.05),观察组患者随着病情的加重,其血清IMA、MCP-1、CD163水平与Gensini评分、血浆NT-proBNP水平呈明显上升趋势;患者血清IMA、MCP-1、CD163这三个指标都是水平越高、Gensini评分越高,血浆NT-proBNP水平越高,呈现正相关;血清IMA、血清MCP-1、血清CD163均为ACS的独立危险因素,而血清HDL为ACS的保护因素。结论 IMA、MCP-1、CD163与NT-proBNP水平和Gensini评分均呈正相关关系,与HDL均是ACS的独立危险因素,有望成为ACS病情评估和预后预测的指标。     

Macrophages expressing the scavenger receptor CD163: a link between immune alterations of the gut and synovial inflammation in spondyloarthropathy

The objective of this study was to investigate CD163+ macrophages in the synovial membrane of patients with spondyloarthropathy (SpA). Immunohistochemistry was performed on synovium of 17 SpA and 18 rheumatoid arthritis (RA) patients, on colonic biopsies of 16 SpA patients and ten healthy controls, and on paired synovial biopsies of eight SpA patients, before and after anti-TNFalpha therapy. Phenotype and cytokine production were analysed by flow cytometry. CD163+ macrophages were increased in the synovial lining and sublining in SpA versus RA, as well as in colonic lamina propria in SpA versus controls. The number of CD163+ macrophages in the synovial sublining correlated with C-reactive protein levels and erythrocyte sedimentation rate. Paralleling the increase of CD163, HLA-DR was increased in the synovial lining and sublining of SpA. In contrast, the co-stimulatory molecules CD80 and CD86 and the dendritic cell markers CD1a and CD83 were scarce in SpA synovium. Flow cytometry indicated that CD163+ macrophages expressed high levels of HLA-DR and could produce in vitro tumour necrosis factor alpha (TNFalpha) but not interleukin-10 (IL-10). Finally, anti-TNFalpha therapy in vivo induced a decrease of CD163+ macrophages in the synovial lining and sublining. In conclusion, macrophages expressing the scavenger receptor CD163 are increased in synovium and in colonic mucosa in SpA, highlighting the relationship between joint and gut in this disease. The correlation with inflammatory parameters, the expression of HLA-DR, the production of TNFalpha but not IL-10, and the reduction by anti-TNFalpha therapy support a role for CD163+ macrophages in the synovial inflammation in SpA.     

Glycosyl-phosphatidylinositol (GPI)-anchored membrane association of the porcine reproductive and respiratory syndrome virus GP4 glycoprotein and its co-localization with CD163 in lipid rafts

The porcine reproductive and respiratory syndrome virus (PRRSV) glycoprotein 4 (GP4) resembles a typical type I membrane protein in its structure but lacks a hydrophilic tail at the C-terminus, suggesting that GP4 may be a lipid-anchored membrane protein. Using the human decay-accelerating factor (DAF; CD55), a known glycosyl-phosphatidylinositol (GPI) lipid-anchored protein, chimeric constructs were made to substitute the GPI-anchor domain of DAF with the putative lipid-anchor domain of GP4, and their membrane association and lipase cleavage were determined in cells. The DAF-GP4 fusion protein was transported to the plasma membrane and was cleaved by phosphatidylinositol-specific phospholipase C (PI-PLC), indicating that the C-terminal domain of GP4 functions as a GPI anchor. Mutational studies for residues adjacent to the GPI modification site and characterization of respective mutant viruses generated from infectious cDNA clones show that the ability of GP4 for membrane association corresponded to virus viability and growth characteristics. The residues T158 (ω − 2, where ω is the GPI moiety at E160), P159 (ω − 1), and M162 (ω + 2) of GP4 were determined to be important for virus replication, with M162 being of particular importance for virus infectivity. The complete removal of the peptide-anchor domain in GP4 resulted in a complete loss of virus infectivity. The depletion of cholesterol from the plasma membrane of cells reduced the virus production, suggesting a role of lipid rafts in PRRSV infection. Remarkably, GP4 was found to co-localize with CD163 in the lipid rafts on the plasma membrane. Since CD163 has been reported as a cellular receptor for PRRSV and GP4 has been shown to interact with this receptor, our data implicates an important role of lipid rafts during entry of the virus.     

Immunohistochemical determination of CD23 expression in Hodgkin's disease using paraffin sections

The leukocyte antigen CD23 is expressed during B-cell development, and functions as an IgE receptor and a lymphocyte growth factor. We studied the expression of CD23 in paraffin sections of lymphoid tissue using the monoclonal antibody BU38. Fifteen cases of Hodgkin's disease, ten reactive lymph nodes, eight B-cell, and seven T-cell non-Hodgkin's lymphomas were analysed immunohistologically. CD23 positivity was seen on follicular dendritic cells and a small number of lymphocytes in reactive nodes. Thirteen of the 15 cases of Hodgkin's disease showed CD23 expression in both neoplastic cells and reactive lymphocytic infiltrate. The antigen was demonstrated in four of the B-cell and one of the T-cell tumours. CD23 may be important in mediating the mixed cellular infiltrate characteristic of Hodgkin's lymphoma.     

CD56 positive diffuse large B-cell lymphoma: a case report and literature review

CD56 positive B-cell lymphoma is very rare. We experienced a case of CD56 positive diffuse large B-cell lymphoma, occurred in a young child. A 5-year-old girl complained with snoring and open mouth breathing. No any abnormality in laboratory or physical examination was present, except enlarged both tonsils. Bilateral tonsillectomy was performed. Cut sections of right tonsil showed a 2 cm size, solid mass. On microscopically, large monomorphic lymphoid cells were diffusely proliferated and showed positivity for CD20 and CD56 and negative for Epstein-Barr virus (EBV) polymerase chain reaction (PCR). Monoclonality was observed on immunoglobulin heavy chain gene rearrangement. This is a unique case with incidentally found and occurred in a young child.     

B cells in the aged: CD27, CD5, and CD40 expression

Ageing is characterized by numerous changes in lymphocyte subpopulations. In the present paper we have focused on B cells carrying the surface markers CD27, CD5 and CD40. CD27 is considered a marker of primed (memory) cells and its engagement promotes the differentiation of memory B cells into plasma cells. CD5 is expressed on B1 cells, which are considered to be responsible for T cell-independent antibody production other than autoantibodies. The CD40 molecule binds CD40L (CD154) and is necessary for T-dependent antibody responses. Here we show that the absolute number of CD5+ and CD40+ B cells is decreased in the elderly, while CD27+ B lymphocytes only marginally decrease in centenarians. However, there is a decrease of the percentage of CD5+ B cells, an increase of CD27+ B cells, while CD40 does not change significantly. These data, together with the increased number of NK cells during aging, suggest different regulation of antibody production in the elderly which might be another example of immune remodeling with aging, based on interactions between human B and NK cells.