First published online June 28, 2004
doi: 10.1242/10.1242/dev.01189
Development 131, 3457-3467 (2004)
Published by The Company of Biologists 2004
orpk mouse model of polycystic kidney disease reveals essential role of primary cilia in pancreatic tissue organization
David A. Cano1,
Noel S. Murcia2,
Gregory J. Pazour3 and
Matthias Hebrok1,*
1 Diabetes Center, Department of Medicine, University of California, San
Francisco, CA 94143, USA
2 Department of Pediatrics, Case Western Reserve University, Cleveland, OH
44106, USA
3 Program in Molecular Medicine, University of Massachusetts Medical School,
Worcester, MA 01605, USA
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Fig. 1. Defects in pancreas maturation in orpk mice. Pancreatic mass is
reduced in orpk mice (B) when compared with wild type (A) at P9. To
adjust for differences in body mass, pancreas weight was divided by body
weight (C; blue, wild type, n=5; red, orpk, n=5).
Progressive acinar cell loss and ductular hyperplasia increase in
orpk mice with age (D,F-H) when compared with wild type (E,I). Gomori
trichrome staining reveals a significant increase in collagen deposition in
orpk pancreas (K) when compared with wild-type tissue (J). d,
duodenum; dp, dorsal pancreas; s, stomach; vp, ventral pancreas.
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Fig. 4. Increase of acinar cell apoptosis and ductal proliferation in orpk
mice after birth (P4). TUNEL staining showed an increase of apoptotic cells in
pancreas of orpk mice (B) when compared with wild-type mice (A).
Inset in B shows that no apoptotic cells could be detected in islets
(outlined) in orpk mice. Proliferation is increased in dilated ducts
of orpk mice, as shown by Ki-67 staining (C,D). Note that the number
of proliferating cells in orpk mice in dilated ducts (outlined in
white) is higher than in non-dilated ducts (outlined in yellow).
Quantification of apoptotic cells, measured as number of cells positive for
TUNEL staining per field of view (E; blue, wild type, n=5; red
orpk, n=5), showed an increase of cell apoptosis in orpk
mice. Proliferation indexes of pancreatic cells are given as number of
Ki-67-positive cells per mucin-positive, insulin-positive or amylase-positive
cells (F; wild type, n=5; orpk, n=5). Wild-type numbers for
apoptosis and proliferation were set to `1' to facilitate comparison (E,F).
Errors bars are shown as ±s.e.m.
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Fig. 2. Tg737 expression in pancreatic tissue and islet formation in
orpk mice. Staining for ß-galactosidase activity in heterozygous
Tg737 mice that carry the lacZ gene under control of the
endogenous Tg737 promoter reveals broad expression in the pancreas at
E15.5 (A). Expression becomes progressively confined to islets and
intercalated ducts in newborn (B) and adult animals (C). Insets in B and C
show expression in ductal cells (arrows). Acini in A and islets in B are
outlined for a better visualization. Islet formation is unaffected in
orpk mice (E,F). Islets were stained with antibodies directed against
centrally located insulin (green) and marginally located glucagon-producing
cells (red). Quantification of islet areas revealed no difference between
wild-type and orpk mice at P9 (D). To adjust for differences in body
mass, islet area was divided by body weight. (D; blue, wild type,
n=5; red, orpk, n=5). Wild-type islet area was adjusted to
`1' to facilitate comparison. Errors bars are shown as ±s.e.m.
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Fig. 3. Acinar cell loss and ductular hyperplasia in orpk mice. Staining
for amylase indicates normal exocrine architecture in E18.5 orpk
pancreatic tissue (A,B). Mucin 1 staining reveals ductal defects in
orpk mice with small dilations of intracinar ducts (arrow in B).
After birth (P4), extensive acinar cell loss occurs, as shown by the decrease
of amylase staining in orpk mice (C,D; shown at higher magnification
in E,F). Increased dilation of duct-like structures was observed after birth
(P4), as shown by mucin staining (C-F). Insulin-expressing cells are present
in duct-like structures of orpk mice at P9 (H, arrows). No
insulin-expressing cells could be detected in ducts of wild-type mice at P9
(G).
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Fig. 5. Absence of pancreatic cilia and increased polycystin-2 expression in
orpk mice. In wild-type pancreatic sections (A) polycystin-2
expression is localized in intercalated ducts. Cilia were visualized using an
antibody against acetylated tubulin. Cilia are confined to the lumen of
intracinar and intercalated ducts (arrows). Acini (a) and intercalated ducts
(id) are outlined in yellow and white, respectively, for better visualization.
Inset shows Hematoxylin/Eosin staining of acini in wild type. A dramatic
decrease in cilia number is detected in orpk mice (B; arrow).
Polycystin-2 expression in orpk mice is increased and mislocalized in
intracinar dilated ducts. An acini with a severe intracinar ductal dilation is
outlined in yellow. Dilated intracinar ducts (outlined in red), a structure
that is not found in wild-type pancreas, show high polycystin-2 expression,
comparable to levels that are only observed in intercalated ducts of control
mice. Inset shows Hematoxylin/Eosin staining of a dilated intracinar duct in
orpk mice. Cilia are also present in islets in wild-type mice (C),
but are almost absent in orpk mice (D; arrow). Cilia are present at
embryonic stages (E15.5) in wild type (E) but very few could be detected in
orpk mice (F). Cilia length, shown in µm, is reduced in
orpk mice (G). Tissue sections were derived from three different
animals at each stage. Errors bars are shown as ±s.d. a, acini; id,
intercalated duct; dd, dilated ducts.
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Fig. 6. Pancreatic abnormalities in inversin and Pkd2 mice.
Hematoxylin/Eosin staining of wild-type (A,C), inversin (B) and Pkd2
mice (D), showing changes in pancreas architecture. Staining for amylase and
mucin indicates normal architecture in wild-type littermates (E,G), but acinar
cell loss and ductal dilation in inversin and Pkd2 mice (F,H).
Arrowheads in H indicate dilated ducts. Elevated levels of acinar cell
apoptosis were observed in inversin mice (J) compared with wild-type
littermates (I). No differences in apoptosis levels were detected between
Pkd2 (L) and wild-type (K) E14.5 embryos.
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Fig. 7. Abnormal expression of ß-catenin and Lef/Tcf family members in
orpk mice. In wild-type pancreatic sections ß-catenin is
predominately localized to the cell membrane (A; arrows in higher
magnification image, C). In orpk mice, cytoplasmic ß-catenin
localization in dilated ducts (arrowheads) is increased (B; and at higher
magnification in D). (E) Expression of Lef/Tcf family members in control
littermates (first lane) and orpk mice (second and third lane). Actin
was used as a control gene.
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© The Company of Biologists Ltd 2004
