Since GCEC injury characterizes the renal manifestations of HUS, understanding the mechanisms of VT-induced GCEC injury is of importance. We previously demonstrated that human GCEC and MC are susceptible to VT-1 and develop altered cell shape and cell death. Cell death may develop through necrosis or programmed cell death (apoptosis). The role of apoptosis in VT-1 mediated glomerular cell injury was investigated in human GCEC and MC in culture.

Pure cultures in serum-replete media were exposed to varying concentrations of VT-1 (1 pM to 1 nM) for 6-48 hr. Cell morphology was assessed by phase contrast microscopy and Giemsa staining. Apoptotic cells were defined by appearance of nuclear condensation and cytoplasmic contraction. Phalloidin was used to define cytoskeletal F-actin. Nucleosomal DNA fragments were determined by TdT in situ apoptosis detection.

GCEC and MC morphology was altered after 6 hr VT-1 with irregular cell spreading and borders noted. GCEC apoptosis was detected after 6 hr 10 pM VT-1(6.6%) and 24 hr 10 pM VT-1 (8%) compared to 1% in control cells. MC apoptosis was also noted with VT-1. In situ TdT labeling revealed nuclear changes consistent with apoptosis at each time period after VT-1. GCEC and MC F-actin was altered after 6 hr VT-1 with more pronounced alterations after 24 hr exposure.

Human GCEC and MC are susceptible to VT-1 and undergo apoptosis in association with cell detachment and cell loss. The initiation of apoptosis after interaction with VT-1 may explain some of the glomerular cell changes seen in HUS. Further understanding of GCEC apoptosis may offer opportunities for intervention to prevent progressive glomerular injury in children with HUS.