Pdx-1, an important transcription factor highlighting in the early pancreatic development, islet functions and pancreatic disorders, needs to be more investigated in zebrafish, and siRNA is still seldom applied in zebrafish embryo-related research. Our aim was to explore the role of pdx-1 in pancreatic development of zebrafish embryos by using siRNA approach. Microinjection, reverse transcriptase-PCR (RT-PCR), in situ hybridization and immunofluorescent staining were used in this research, and the morphology of the islet in normal zebrafish embryos, and in those treated with the siRNA specific to pdx-1 (siPDX-1) or siGFP was observed and compared. The expression of pdx-1 was detected in the stages of 1-cell, 2-cell, 4-cell, 8-cell, 16-cell, 16-hour by RT-PCT. The in situ hybridization and immunofluorescent staining results showed that siPDX-1 disturbed the formation of the islet in zebrafish embryos. Pdx-1 played multiple roles in maintaining the phenotype of the islet during embryogenesis in zebrafish.
Embryo, Nonmammalian ; Homeodomain Proteins/genetics ; Homeodomain Proteins/*metabolism ; Islets of Langerhans/cytology ; Islets of Langerhans/*embryology ; Islets of Langerhans/metabolism ; RNA Interference ; RNA, Small Interfering/*genetics ; Trans-Activators/genetics ; Trans-Activators/*metabolism ; Zebrafish

Pancreas islet cell transplantation has been regarded as an ideal method to treat the type I diabetes mellitus. However, it could not be the method of choice because of poor graft survival rate after transplantation. Recently, the outcome of pancreas islet cell transplantation has been improving, especially since the Edmonton group has succeeded in controlling the glucose metabolism in 7 consecutive type I diabetes mellitus patients. Returning to diabetic status in a substantial portion of transplanted patients, however, has revealed that lots of hurdles, such as primary non-function of the islet from non-specific inflammation, immunologic destruction of islets from either allogenic or autoimmune process, and shortage of donor source, remained to be solved in the near future, if pancreas islet cell transplantation is to be a practical clinical treatment modality for diabetic patients. We herein discuss on the current status and future of pancreas islet cell transplantation.
Diabetes Mellitus ; Glucose ; Graft Survival ; Humans ; Inflammation ; Islets of Langerhans* ; Metabolism ; Pancreas* ; Tissue Donors ; Transplantation

Insulin is produced and secreted by pancreatic β cells in the pancreas, which plays a key role in maintaining euglycemia. Insufficient secretion or deficient usage of insulin is the main cause of diabetes mellitus (DM). Drug therapy and islets transplantation are classical treatments for DM. Pancreatic β cell replacement therapy could help patients to get rid of drugs and alleviate the problem of lacking in transplantable donors. Pancreatic β-like cells can be acquired by cell reprogramming techniques or directed induction of stem cell differentiation. These cells are proved to be functional both in vitro and in vivo. Some hospitals have already performed clinical trials for pancreatic β cell replacement therapy. Functional pancreatic β-like cells, which obtained from in vitro pathway, could be a reliable source of cell therapy for treating DM. In this review, the approaches of obtaining pancreatic β cells are summarized and the remaining problems are discussed. Some thoughts are provided for further acquisition and application of pancreatic β cells.
Cell Differentiation ; Diabetes Mellitus/therapy* ; Humans ; Insulin/metabolism* ; Insulin-Secreting Cells/metabolism* ; Islets of Langerhans Transplantation ; Pancreas/metabolism*

BACKGROUNDThe induced expression of heme oxygenase-1 (HO-1) in donor islets improves allograft survival. Cobalt protoporphyrin (CoPP) could significantly enhance the expression of HO-1 mRNA and protein in rat islet safely. Our work was to study how to protect pancreatic islet xenograft by CoPP-induction.

METHODSIslet xenografts treated with CoPP-induction and CoPP + Zinc protoporphyrin (ZnPP) in vitro and in vivo were randomly transplanted into murine subrenal capsule; then the graft survival time was compared by blood glucose level and pathological examination and meanwhile the interferon γ (IFN-γ), tumor necrosis factor α (TNF-α), interleukin 10 (IL-10) and IL-1β level in serum and their mRNA and HO-1 mRNA and protein expression were examined.

RESULTSIslets with CoPP-induction under low- and high-glucose stimulation exhibited much higher insulin secretion compared with other three groups. CoPP-induction could increase higher expression of HO-1 (mRNA: 3.33- and 76.09-fold in vitro and in vivo; protein: 2.85- and 58.72-fold). The normoglycemia time in induction groups ((14.63 ± 1.19) and (16.88 ± 1.64) days) was significantly longer. The pathological examination showed less lymphocyte infiltration in induction groups. The IL-10 level and its mRNA in induction groups were significantly higher.

CONCLUSIONSThe HO-1 induced by CoPP would significantly improve function, prolong normoglycemia time and reduce lymphocyte infiltration. Meanwhile CoPP-induction in vivo had more beneficial effects than in vitro. Its mechanism could be related to immune-modulation of IL-10.

Animals ; Blotting, Western ; Graft Survival ; Heme Oxygenase-1 ; genetics ; metabolism ; Interleukin-10 ; blood ; Islets of Langerhans ; enzymology ; metabolism ; Islets of Langerhans Transplantation ; Male ; Mice ; Mice, Inbred BALB C ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Transplantation, Heterologous

Alzheimer Disease ; metabolism ; Animals ; Brain ; metabolism ; Calcium-Binding Proteins ; biosynthesis ; chemistry ; genetics ; Gastrointestinal Tract ; metabolism ; Humans ; Islets of Langerhans ; metabolism ; Neoplasms ; metabolism ; RNA, Messenger ; metabolism ; Secretagogins ; Thyroid Gland ; metabolism

The regional distribution and relative frequency of the pancreatic endocrine cells in the C57BL/6 mouse were studied by immunohistochemical method using four types of specific mammalian antisera against insulin, glucagon, somatostatin and human pancreatic polypeptide (PP). The pancreas of mouse could be divided into three portions; pancreatic islets, pancreatic duct and exocrine portions, and pancreatic islets were further subdivided into three regions (central, mantle and peripheral regions) according to their located types of immunoreactive cells and pancreatic duct portions were also subdivided into two regions (epithelial and connective tissue regions). In the pancreatic islet portions, although some cells were also demonstrated in the mantle regions, most of insulin-immunoreactive cells were located in the central regions and they were randomly dispersed in the whole pancreatic islets. Glucagon-immunoreactive cells were detected in the mantle and peripheral regions. Their relative frequencies in the peripheral regions were somewhat numerous than those of the mantle regions. Somatostatin-immunoreactive cells were detected in the mantle and peripheral regions. However, no PP-immunoreactive cells were demonstrated in the pancreatic islets of C57BL/6 mouse. In the pancreatic duct portions, rare glucagon-immunoreactive cells were situated in the epithelial regions. Cell clusters that consisted of glucagon- or somatostatin- immunoreactive cells were found in some case of connective tissue regions of pancreatic ducts. However, insulin- and PP-immunoreactive cells were not detected in the epithelial nor connective tissue regions. In the exocrine portions, all four types of immunoreactive cells except for PP cells were demonstrated in the C57BL/6 mouse. However, no PP-immunoreactive cells were demonstrated. In conclusion, regional distribution of endocrine cells in the pancreas of C57BL/6 mouse was similar to that of mammals, especially other rodents except for topographically different distribution of endocrine cells compared to that of other rodents.
Animals ; Female ; Glucagon/metabolism ; Immunohistochemistry ; Insulin/metabolism ; Islets of Langerhans/cytology/*metabolism ; Male ; Mice ; Mice, Inbred C57BL/*metabolism ; Pancreatic Polypeptide/metabolism ; Somatostatin/metabolism

The regional distribution and relative frequency of insulin-, glucagon-, somatostatin- and pancreatic polypeptide (PP)-producing endocrine cells in the pancreas of BALB/c mouse were investigated by immunohistochemical method. The pancreas of mice was divided into two portions; pancreatic islets and exocrine portions, and pancreatic islets were further subdivided into two regions (central and peripheral regions) and the relative frequency and regional distribution of immunoreactive cells against insulin, glucagon, somatostatin and PP antisera were monitored. In the pancreatic islet portions, insulin-immunoreactive cells were located in the central regions and they were randomly dispersed in the whole pancreatic islets in some case of the small islets. Quite different from those of other mammals, glucagon-immunoreactive cells were dispersed throughout central to peripheral regions in case of large islets and in the smaller ones, most of these cells were situated in the peripheral regions. Somatostatin-immunoreactive cells were detected in the peripheral regions with various frequencies. Although some cells were demonstrated in the central regions of pancreatic islets, most of PP-immunoreactive cells were located in the peripheral regions. In the exocrine portions, all four types of immunoreactive cells were demonstrated in the BALB/c mouse. Some peculiar distributional patterns of pancreatic endocrine cells were found in BALB/c mouse, especially in case of glucagon-immunoreactive cells.
Animals ; Female ; Glucagon/*metabolism ; Immunohistochemistry/veterinary ; Insulin/metabolism ; Islets of Langerhans/cytology/*metabolism ; Male ; Mice ; Mice, Inbred BALB C/*metabolism ; Pancreatic Polypeptide/metabolism ; Somatostatin/metabolism

PURPOSE: Transplantation of pancreas islet has been worldwidely studied as a one of therapeutic modalities to achieve the insulin independence. We studied whether the expression of vascular endothelial growth factor (VEGF) on pancreas islets with liposomal VEGF gene transfer could improve the efficacy of early implantation and long term graft survival in pancreatic islet cell transplantation. METHODS: Syngenic pancreas islets were transplanted beneath the renal capsule. Islets were transfected with plasmid VEGF c-DNA using cationic liposome DMRIE-C. Glucose metabolism and histologic findings were compared between the groups transplanted with VEGF DNA containing islets (n=5) and the control group with (n=5) or without (n=4) local recombinant VEGF adminstration during islet transplant. RESULTS: Glucose was controlled at 5.5 days after transplantation in control group without r-VEGF adminstration, at 4 days in group with recombinant VEGF adminstration, and at 6.6 days in group with VEGF DNA transfected islets. Euglycemia was maintained over 150 days in control group. However, graft failure was developed in 22 days after transplantation in group with VEGF DNA transfected islet. Histologically there were severe infiltrations of neutrophil and lymphocyte in VEGF DNA transfected grafts from 5 days after transplantation. CONCLUSION: Although VEGF could be a favorable angiogenic factor in pancreas islet transplantation, VEGF expression following VEGF DNA transfection into islets could not increase the graft survival due to inflammatory process. More investigations are needed to clarify the mechanism on destructive process of islets after gene transfection into islets, and another approaches to get the effect of gene transfection should be followed.
Angiogenesis Inducing Agents ; Cell Transplantation* ; DNA ; Endothelial Growth Factors* ; Glucose ; Graft Survival* ; Insulin ; Islets of Langerhans Transplantation ; Islets of Langerhans* ; Liposomes ; Lymphocytes ; Metabolism ; Neutrophils ; Pancreas ; Plasmids ; Transfection* ; Transplants* ; Vascular Endothelial Growth Factor A

OBJECTIVETo explore the synergistic protective effect of co-transplanted testicular cells expressing FasL and CsA on survival of islet allografts.

METHODSThe allogeneic islets and testicular cells were co-transplanted into the renal subcapsular space of the diabetic recipients with or without CsA after operation. Allografts survival period and the testicular cells or islets function were analyzed.

RESULTSThe mean survival period of control group was 4.6 +/- 1.1 days. When CsA was administered after transplantation, the mean survival period of islet allografts, (21.8 +/- 4.7) days, was significantly longer than that of control group (P < 0.01). When islets were co-transplanted together with 1 x 10(7) testicular cells (group A), a significant prolongation of graft survival was found (more than 57.5 +/- 4.0 days; P < 0.01 vs. control). But if 1 x 10(7) testicular cells expressing FasL were cultured with FasL-mAb for 30 minutes before co-transplantation (group B), the mean survival period of islet allografts (5.8 +/- 2.6 days), was similar to that in control group, but significantly shorter than that in group A (P < 0.01). When islets and 1 x 10(5) testicular cells were co-transplanted separately into the bilateral renal subcapsular space with CsA (group C), the survival of islet allografts was significantly prolonged in comparison with control group (more than 55.0 +/- 6.5 days; P < 0.01 vs. control), and similar to islets co-transplanted together with 1 x 10(7) testicular cells (group A). When islets were co-transplanted separately with 1 x 10(6) testicular cells without CsA (group D), the mean survival period (11.5 +/- 3.1 days) was shorter than that in group C, but prolonged in comparison to control group (P < 0.05).

CONCLUSIONThe co-transplanted testicular cells expressing FasL with administering CsA post-transplantation can jointly inhibit immune rejection of islet allografts by different mechanism and play a systemic and synergistic protective role to islet allografts.

Animals ; Cyclosporine ; therapeutic use ; Fas Ligand Protein ; Graft Survival ; Immunohistochemistry ; Insulin ; blood ; Islets of Langerhans ; pathology ; ultrastructure ; Islets of Langerhans Transplantation ; Male ; Membrane Glycoproteins ; analysis ; genetics ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Sertoli Cells ; metabolism ; transplantation ; Transplantation, Homologous

This study was performed to observe the changes of glucose-related hormones and the morphological change including ultrastructure of the pancreatic islets in the male Otsuka Long-Evans Tokushima Fatty rat. Area under the curve (AUC) of glucose at the 30th (709 +/- 73 mg.h/dL) and at the 40th week (746 +/- 87 mg.h/ dL) of age were significantly higher than that at the 10th week (360 +/- 25 mg.h/ dL). AUC of insulin of the 10th week was 2.4 +/- 0.9 ng.h/mL, increased gradually to 10.8 +/- 8.3 ng.h/mL at the 30th week, and decreased to 1.8 +/- 1.2 ng.h/mL at the 40th week. The size of islet was increased at 20th week of age and the distribution of peripheral alpha cells and central beta cells at the 10th and 20th weeks was changed to a mixed pattern at the 40th week. On electron microscopic examination, beta cells at the 20th week showed many immature secretory granules, increased mitochondria, and hypertrophied Golgi complex and endoplasmic reticulum. At the 40th week, beta cell contained scanty intracellular organelles and secretory granules and apoptosis of acinar cell was observed. In conclusion, as diabetes progressed, increased secretion of insulin was accompanied by increases in size of islets and number of beta-cells in male OLETF rats showing obese type 2 diabetes. However, these compensatory changes could not overcome the requirement of insulin according to the continuous hyperglycemia after development of diabetes.
Animals ; Body Weight ; Diabetes Mellitus, Type 2/*metabolism/pathology ; Disease Models, Animal ; Glucagon/*metabolism ; Insulin/*metabolism ; Islets of Langerhans/*metabolism/pathology/ultrastructure ; Male ; Rats ; Rats, Inbred OLETF