Proapoptotic Bax is hyperexpressed in isolated human islets compared with antiapoptotic Bcl-2.
BACKGROUND
Apoptosis is a well-documented pathway for islet cell death. One potential mechanism is overexpression of death-promoting Bax compared with antiapoptotic Bcl-2 in islets.


METHODS
We isolated islets from 10 human pancreata and measured the expression of Bax mRNA and Bcl-2 mRNA by real-time quantitative polymerase chain reaction; islet and pancreas expression of Bax, Bcl-2, activated caspase-3, and cleaved poly (ADP-ribose) polymerase were also assessed by immunohistochemistry. Islet cell apoptosis was evaluated by terminal deoxynucleotide transferase-mediated dUTP nick-end labeling (TUNEL) assay and by flow cytometry.


RESULTS
The mean (+/-SE) level of Bax mRNA was 336+/-79 copies per nanogram of total RNA, and the level of Bcl-2 mRNA was 36+/-10 (P=0.001). A positive correlation existed between islet expression of Bax mRNA and Bcl-2 mRNA (P=0.001). The islet Bax to Bcl-2 ratio was 10.8+/-1.3 and 1.71+/-0.3 for the spleens (P=0.0001). Bax mRNA (P=0.04), but not Bcl-2 mRNA, was expressed at a higher level in islets compared with spleens. Human islets contained large numbers of cells expressing Bax protein, whereas only infrequent islet cells expressed Bcl-2 protein, activated caspase-3, and poly (ADP-ribose) polymerase. The apoptotic index was 5% by TUNEL assay, and the percentage of apoptotic islet cells was 9.7+/-2.5% by flow cytometry. Sections of human pancreas before islet isolation showed islet staining for Bax but not Bcl-2.


CONCLUSIONS
Our finding that isolated human islets express Bax at a higher level compared with Bcl-2 suggests a molecular mechanism for islet cell death by apoptosis. We hypothesize that reducing islet expression of Bax, or regulating its activation, will help preserve islet cell mass after islet transplantation.
Thomas
Dolca
D
Division of Nephrology, Department of Medicine, Weill Medical College of Cornell University, New York, NY, USA.
Yang
Hua
H
Boffa
Daniel J
DJ
Ding
Ruchuang
R
Sharma
Vijay K
VK
Lagman
Milagros
M
Li
Baogui
B
Hering
Bernhard
B
Mohanakumar
Thalachallour
T
Lakey
Jonathan
J
Kapur
Sandip
S
Hancock
Wayne W
WW
Suthanthiran
Manikkam
M
D000328:Adult; D017209:Apoptosis / Q000502:physiology*; D015925:Cryopreservation; D005260:Female; D006801:Humans; D007515:Islets of Langerhans / Q000378:metabolism* / Q000502:physiology; D008297:Male; D008875:Middle Aged; D010179:Pancreas; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000235:genetics / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D013997:Time Factors; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein