Long-term follow-up failed to detect in vitro transmission of full-length porcine endogenous retroviruses from specific pathogen-free pig islets to human cells

Abstract

Aims/hypothesis. Islets from specific pathogen-free (SPF) pigs could prevent the transmission of conventional zoonosis, but not endogenous retroviruses (PERV), from pigs to diabetic patients. We previously reported that the pancreas showed the lowest expression of PERV mRNA among pig tissues intended for grafting. This study aimed to determine whether PERV from pig islets infect human cells during co-incubation. Methods. Human cells (including highly PERV-sensitive 293 cells) were incubated with SPF pig islet cells under conditions designed to increase contact (a high islet to human cell ratio, extended period of coculture, and repeated contacts). PK15 and G2 retrovirus-producing pig cells were used in place of islet cells as ªpositive infection controlsº. Infection of human cells was monitored on cellular extracts and supernatants by PCR or long PCR, and RT-PCR or long RT-PCR, to detect PERV DNA and mRNA, respectively. Reverse-transcriptase activity was monitored by PERT. Results. Despite the presence of all PERV sequences in pig islet cells, including full-length inserts, no DNA or RNA for gag, pol, and the 3 env sub-types were detected in any human cell line or blood mononuclear cells incubated with pig islet cells, during an 18-week follow-up period. No PERV sequences or RT activity were detected in supernatants. PERV signals were negative even when the pig islet to human cell ratio was increased to 100:1, the time of co-culture was extended to 5 days and two sequential co-incubations were done. By contrast, all PERV DNA and mRNA were detected in all human cells co-incubated with PK15 or G2 cells. Depending on human cell types, productive or non-productive infections were obtained: full-length PERV RNA and RT activity in supernatants were detected or not; and PERV sequences to previously unexposed human cells by PERV-infected human cells were transmitted or not. Some human cells were not productively infected by PK15 cells but became productively infected after co-incubation with PERV-infected 293 cells. Conclusion/interpretation. SPF pig islet cells, even with PERV inserts and transcripts, have very little probability of transmitting PERV to human cells during co-incubation. The sensitivity of human cells to stable and productive infection by PERV depends on the cell type. Human adaptation of PERV was observed. [Diabetologia (2001

Microchimerism and transmission of porcine endogenous retrovirus from a pig cell line or specific pathogen-free pig islets to mouse tissues and human cells during xenografts in nude mice

“…1), but no full-length PERV RNA, were found in pig islet cells (n=8). As already reported by us, the expression of all PERV mRNA was much lower in pig islet cells than in PK15 cells [5,8].…”

“…The primers used for PCR amplification of PERV sequences or pig mt DNA were synthesised by the Life Tech-During xenograft, there is a risk of transmitting infectious agents from pig to man. The risk of conventional zoonosis led us to isolate islets from specific pathogen-free (SPF) pigs [1,2,3,4,5,6,7,8]. However, there is also a risk of transmitting porcine endogenous retroviruses (PERV) [9,10,11,12].…”

“…However, there is also a risk of transmitting porcine endogenous retroviruses (PERV) [9,10,11,12]. The ability of PERV [10] produced by pig cells to infect human cells in vitro has been documented [8,9,13,14,15,16,17]. PERV mRNA and full-length endogenous retroviral genome were expressed in all pigs tested [9,15,18,19], including SPF pigs [5,20].…”

“…The risk of transmission could depend on viral load, as in the case of other retroviral infections [34]. Our previous work showed that pancreas expressed the lowest levels of PERV mRNA in SPF pig tissues intended for grafting [5], and that long-term follow-up failed to detect in vitro transmission of PERV from SPF pig islets to a panel of human cells, even after repeated and optimised co-incubations [8].…”

“…Full-length PERV RNA (long RT-PCR), PERV mRNA (RT-PCR) and RT activity (PERT) were detected in supernatants of PK15 cells (n=12) [5,8]. In contrast, full-length PERV transcripts and RT activity were never detected in supernatants of SPF pig islet cells (n=10).…”