感染猫白血病病毒的猫中传染性逆转录病毒在软骨中的定位。

Arnoczky Steven P, Swenson Cheryl, Egerbacher Monika, Gardner Keri, Caballero Oscar, Burns Meghan

Laboratory for Comparative Orthopaedic Research and Department of Pathobiology and Diagnostic Investigation, College of Veterinary Medicine, Michigan State University, East Lansing, MI 48824, USA.

J Bone Joint Surg Am. 2007 Sep;89(9):2030-6. doi: 10.2106/JBJS.G.00054.

BACKGROUND

Previous studies have suggested that articular cartilage allografts were not likely to transmit infectious retrovirus since viral DNA could not be isolated from chondrocytes of infected individuals. However, the ability of the extracellular matrix of articular cartilage to harbor and transmit a retrovirus has not been examined. We hypothesized that articular cartilage fragments, but not isolated chondrocytes, from cats systemically infected with feline leukemia virus (FeLV) are capable of transmitting infectious retrovirus.

METHODS

Fresh cartilage segments and chondrocytes isolated from cats systemically infected with feline leukemia virus were used in this study. Feline embryonic fibroblast cells were cocultured with segments of cartilage, isolated chondrocytes, or fragments of cortical bone from each infected cat. The FeLV p27 antigen was measured in the coculture media by enzyme-linked immunosorbent assay. In addition, FeLV proviral nucleic acids were quantified by real-time quantitative polymerase chain reaction with use of DNA extracted from feline embryonic fibroblast cell cocultures as well as isolated chondrocytes. Immunohistochemistry was used to assess for FeLV p27 antigen in both intact cartilage fragments and isolated chondrocytes.

RESULTS

Feline embryonic fibroblast cells cocultured with cartilage fragments from each of the five FeLV-infected cats all demonstrated high levels of proviral DNA, indicating transmission of infective virus. In addition, media from all cocultures of feline embryonic fibroblast cells and chondral fragments became positive for p27 antigen, indicating active viral replication. In contrast, cocultures of feline embryonic fibroblast cells and isolated chondrocytes from all FeLV-infected cats were negative for proviral DNA and p27 antigen. Likewise, no proviral nucleic acids could be detected in isolated chondrocytes from any infected cats. Cocultures of feline embryonic fibroblast cells with cortical bone fragments were positive for proviral DNA and p27 antigen. Immunohistochemical staining of cartilage fragments from FeLV-infected cats demonstrated the presence of p27 antigen throughout the extracellular matrix, but the p27 antigen was not detected in isolated chondrocytes.

CONCLUSIONS

Articular cartilage fragments can readily transmit infectious retrovirus, but isolated chondrocytes were likely not the source of the infectious virus because they did not harbor proviral DNA or p27 antigen.

背景

以往研究表明，同种异体关节软骨不太可能传播感染性逆转录病毒，因为无法从受感染个体的软骨细胞中分离出病毒DNA。然而，关节软骨的细胞外基质容纳和传播逆转录病毒的能力尚未得到研究。我们推测，来自全身感染猫白血病病毒（FeLV）的猫的关节软骨碎片而非分离的软骨细胞能够传播感染性逆转录病毒。

方法

本研究使用从全身感染猫白血病病毒的猫身上分离的新鲜软骨段和软骨细胞。将猫胚胎成纤维细胞与每只受感染猫的软骨段、分离的软骨细胞或皮质骨碎片共培养。通过酶联免疫吸附测定法在共培养培养基中测量FeLV p27抗原。此外，使用从猫胚胎成纤维细胞共培养物以及分离的软骨细胞中提取的DNA，通过实时定量聚合酶链反应对FeLV前病毒核酸进行定量。免疫组织化学用于评估完整软骨碎片和分离的软骨细胞中的FeLV p27抗原。

结果

与五只感染FeLV的猫中的每只猫的软骨碎片共培养的猫胚胎成纤维细胞均显示出高水平的前病毒DNA，表明感染性病毒的传播。此外，猫胚胎成纤维细胞与软骨碎片的所有共培养物的培养基中p27抗原均呈阳性，表明病毒活跃复制。相比之下，所有感染FeLV的猫的猫胚胎成纤维细胞与分离的软骨细胞的共培养物中前病毒DNA和p27抗原均为阴性。同样，在任何受感染猫的分离软骨细胞中均未检测到前病毒核酸。猫胚胎成纤维细胞与皮质骨碎片的共培养物中前病毒DNA和p27抗原呈阳性。对感染FeLV的猫的软骨碎片进行免疫组织化学染色显示，整个细胞外基质中均存在p27抗原，但在分离的软骨细胞中未检测到p27抗原。