Primary differentiated respiratory epithelial cells respond to apical measles virus infection by shedding multinucleated giant cells

Measles virus (MeV) is highly infectious by the respiratory route and remains an important cause of childhood mortality. However, the process by which MeV infection is efficiently established in the respiratory tract is controversial with suggestions that respiratory epithelial cells are not susceptible to infection from the apical mucosal surface. Therefore, it has been hypothesized that infection is initiated in lung macrophages or dendritic cells and that epithelial infection is subsequently established through the basolateral surface by infected lymphocytes. To better understand the process of respiratory tract initiation of MeV infection, primary differentiated respiratory epithelial cell cultures were established from rhesus macaque tracheal and nasal tissues. Infection of these cultures with MeV from the apical surface was more efficient than from the basolateral surface with shedding of viable MeV-producing multinucleated giant cell (MGC) syncytia from the surface. Despite presence of MGCs and infectious virus in supernatant fluids after apical infection, infected cells were not detected in the adherent epithelial sheet and transepithelial electrical resistance was maintained. After infection from the basolateral surface, epithelial damage and large clusters of MeV-positive cells were observed. Treatment with fusion inhibitory peptides showed that MeV production after apical infection was not dependent on infection of the basolateral surface. These results are consistent with the hypothesis that MeV infection is initiated by apical infection of respiratory epithelial cells with subsequent infection of lymphoid tissue and systemic spread.

Measles virus (MeV) is a highly contagious enveloped, nonsegmented negative-strand RNA virus that is transmitted by the respiratory route to cause a systemic rash disease that includes pulmonary infection in both humans and nonhuman primates. Despite availability of an effective live attenuated MeV vaccine (LAMV), the worldwide incidence of measles cases and deaths has recently tripled (1, 2). Because of the efficient respiratory spread of the virus, a major hurdle for control of measles is the requirement for high vaccine-induced population immunity (92 to 95%) to interrupt transmission (3, 4).Although the respiratory tract is the site of wild-type (WT) MeV initiation of infection and aerosol delivery of LAMV induces protective immunity (5), the biology of how MeV so efficiently initiates infection after aerosol or respiratory droplet exposure is controversial. Two fundamentally different models of MeV pathogenesis have been proposed: 1) Initial replication of MeV occurs in respiratory epithelial cells through apical infection with subsequent systemic spread to other cells and tissues; 2) Initial replication of MeV occurs in myeloid cells of the respiratory tract (alveolar macrophages/dendritic cells) with spread to other tissues and subsequent delivery by infected lymphocytes to the basolateral surface of respiratory epithelial cells for pulmonary infection and virus transmission (6, 7). The first model is supported by the presence of giant cells in broncho-alveolar lavage fluid from measles patients and identification of epithelial cell infection by histology or immunohistochemistry in the airway and lung during measles in humans and macaques (8–10). In addition, LAMV can efficiently establish pulmonary infection without producing a viremia (11), suggesting that prior infection of lymphocytes is not necessary for respiratory epithelial cell infection. The alternative model is suggested by the observation that, after pulmonary infection of macaques with recombinant MeV-expressing eGFP, the predominant eGFP⁺ cells in the respiratory epithelium were CD150⁺ cells and dendritic cells, not epithelial cells (12, 13). That epithelial cells are not the site of initial infection is further supported by the apparent resistance of polarized epithelial cells derived from airway tissue to infection with WT MeV via the apical surface (14, 15). Questions remain for both models.Entry of MeV into susceptible cells is determined mainly by interaction of the hemagglutinin (H) and fusion (F) virion surface glycoproteins with receptor molecules on the host cell. The three identified cellular receptors for MeV are CD46 expressed ubiquitously in all nucleated cells (16, 17), CD150 (signaling lymphocyte activation molecule/SLAM) expressed on activated immune cells (18), and nectin-4 expressed by epithelial cells (19, 20). WT MeV can use CD150 and nectin-4 as receptors while LAMVs can utilize all three receptors. Nevertheless, in vitro and in vivo spread of LAMV is more restricted than that of WT MeV (21).The identification of nectin-4 as a functional receptor for MeV on epithelial cells (19, 20) has shed light on the pathogenesis of MeV in the respiratory tract. As an adherens junction protein, nectin-4 is located at the basolateral surfaces of epithelial cells and thus can be used by MeV to spread back into the airways after replicating systemically (22). The importance of nectin-4 in MeV transmission is highlighted by infection of macaques with a recombinant virus unable to recognize nectin-4. This nectin-4-blind virus is defective in shedding virus into the airways, although most other aspects of the infection are indistinguishable from infection caused by WT MeV (7). The observed competency of the nectin-4-blind MeV to enter and establish infection in macaques has been viewed as supporting evidence for the alternative model of MeV pathogenesis in the respiratory tract (20, 23–25). However, the mode of MeV entry into epithelial cells in vivo remains in question and the fact that nectin-4 is located on the basolateral surface of epithelial cells could also indicate that MeV enters respiratory epithelial cells from the apical surface in a nectin-4-independent manner. In particular, other modes of entry, such as endocytosis and macropinocytosis used by several paramyxoviruses, may be important for MeV entry into primary cells (26–28).To better understand MeV infection of the respiratory tract, we have developed and characterized a primary differentiated respiratory epithelial cell culture system using tracheal and nasal epithelial cells derived from rhesus macaques (rmTEC and rmNEC), a highly relevant animal model for measles (29, 30), and compared the outcome of apical and basolateral exposure of these cells to WT MeV and LAMV. Infection occurred through both the apical surface and the basolateral surface. Apical infection was efficient and induced rapid shedding of viable MeV-infected multinucleated giant cells (MGCs) from the epithelial surface into the lumen while maintaining epithelial barrier integrity. Basal infection was less efficient with infected cells retained in the epithelial monolayer and with barrier disruption, as well as with shedding of MGCs from the apical surface.     

Outcome of colonic fistula surgery in the modern surgical era

Background

Various conditions lead to the development of colonic fistulas. Contemporary surgical data is scarce and it is unclear whether advances in surgical care have impacted outcome. The aim of the present study was to review the short- and long-term outcome of patients treated surgically for colonic fistula over an 8-year period at a tertiary institution.

Methods

A retrospective review was performed, focusing on the type of operative interventions, short- and long-term complications, length of hospital stay, readmission rate, mortality rate, and fistula recurrence.

Results

Forty-five patients were treated for colonic fistula. The most common etiology was diverticulitis (74 %). Fistula type was colovesical (58 %), colocutaneous (18 %) and colovaginal (15 %). Laparoscopic resection was performed in 42 % of cases. An intraoperative complication occurred in 4 %. A primary anastomosis was performed in 96 % of patients and 10 (23 %) had a temporary stoma. Median length of hospital stay was 6 days. Postoperative complications were common (47 %) and wound infection was noted in 20 % of patients. The readmission rate was 29 % and the 90-day mortality was 4 %. All patients healed their fistula with no recurrences noted during a median follow-up of 37 months.

Conclusions

Surgical intervention healed the majority of patients with colonic fistula. However postoperative complications were common and readmission occurred in one-third of the cases. Laparoscopic excision was feasible in nearly half of the patients.