Oncogene expression on the syngeneic β-cells of long-term surviving pancreatic grafts and better effects of interleukin-1 receptor (IL-1R) and IL-2Rβ on the grafted β-cells in LEW/Sea strain rats

Abstract

Thirty-two normal LEW/Sea rats were transplanted a piece of syngeneic pancreas between the peritoneum and abdominal muscle. Among them, 17 (68%) of the 25 rats that received pancreatic transplantation at 41–50 days of age had a surviving β-cell mass at 5.5–7.1 months after transplantation. Among the 25 rats, 12 rats injected with interleukin-1 receptor (IL-1R) and IL-2Rβ peptides at post-transplantation showed better surviving grafts at 5.5 months’ observation. Only 2 (25%) of the other 7 young rats that received a pancreatic graft at 20 days of age had a small mass at 21 days post-transplantation. Flow cytometer (FCM) analyses showed that thymus OX40 + (CD134 +) T-cells were increased up to 37±4% at the graft rejection in the 13 old rats without the IL-R peptide injections. The 7 young rats had 99% of thymus OX40 + T-cells. However, the 12 old rats injected with the IL-R peptides showed suppressed numbers of thymus OX40 + T-cells (8–13±3%). The long-term surviving, but apoptotic, grafted β-cells were stained positively both with anti-insulin monoclonal antibody (mAb) and with anti-c-erbB-2/human epidermal growth factor receptor (HER)-2/neu mAb. Expression of a c-erb family oncogene was shown on the pancreatic graft surviving for 7.1 months. Electron microscopic analysis of the grafted β-cells showed abnormally large β granules and loss of functioning mitochondria in the cytoplasm. In 18 (56%) of the 32 rats, the 220-bp and 380-bp specific products of insulin-degrading enzyme (IDE) gene were amplified using the polymerase chain reaction (PCR) of the liver DNA. Among the 18 rats, 6 rats expressed 2 extra hands of 280-bp and 700-bp in a correlation with the high levels of the transforming growth factor-alpha (TGF-α) cDNA of 120-bp which was amplified in the quantitative reverse-transcriptase (RT)-PCR of the liver cDNA. Among the selected 11 rats, 5 rats showed large amounts of the 120-bp TGF-α cDNA. Host pancreatic RT-PCR showed 235-bp or 250-bp bcl-2 and 181-bp bcl-xS gene products. The bcl-2 cDNA of the host pancreas was amplified actively when the pancreatic graft was being rejected. Exceptionally, the one female injected with the IL-R peptides showed a low level of the liver TGF-α cDNA together with the pancreatic expressions of Bax (140-bp), bcl-2 and like interleukin converting enzyme (LICE) (318-bp) cDNA. Insulin secretion from the grafted β-cells and IL-1β-induced Fas-mediated apoptosis of the β-cells were suspected to be present at the same time in the female with the best graft survival.