OBJECTIVETo examine the renal function changes and mechanisms on rats with diabetes through a sleeve gastrectomy operation.

METHODSThirty-six rats were induced diabetes through injection of streptozotocin (STZ), and 30 of these diabetic rats that blood glucose levels at the midrange (blood sugar 17.88-23.65 mmol/L, mean: 20.32 mmol/L) were randomly assigned to the sleeve gastrectomy group, Sham-operation group and control group. The serum creatinine, lipid parameters were measured postoperatively. The 24 h urine volume obtained and urine albumin excretion rate (UAER) was calculated. Serum and urinary creatinine were examined and glomerular filtration rate (GFR) was counted. Kidney sections were stained with periodic acid-Schiff, and then the index of mesangial expansion was determined. The expression of synaptopodin for podocytes was also performed through the immunohistochemical procedure. A one-way ANOVA and t-test were performed to evaluate differences between groups and each other.

RESULTSOnly one rat of SG group died after operation. The GFR ((8.44 ± 2.10) ml · g⁻¹ · d⁻¹), 24 h UAER ((36.04 ± 11.10) mg/d), plasma lipids level (total cholesterol (1.66 ± 0.23) mmol/L, triglycerides (1.25 ± 0.17) mmol/L), kidney weight ((1.61 ± 0.06) g), the index of mesangial expansion ((6.14 ± 1.50)%) and synaptopodin expression ((20.44 ± 2.99)%) were improved in the SG group compared with the sham-operation group ((15.05 ± 3.01) ml · g⁻¹ · d⁻¹, (57.01 ± 11.34) mg/d, (2.15 ± 0.29) mmol/L, (1.65 ± 0.23) mmol/L, (1.93 ± 0.07) g, (11.32 ± 2.09)%, (10.34 ± 1.43)%) and control group ((14.79 ± 2.38) ml · g⁻¹ · d⁻¹, (62.71 ± 16.46) mg/d, (2.23 ± 0.21) mmol/L, (1.59 ± 0.20) mmol/L, (1.91 ± 0.06) g, (10.82 ± 1.79)%, (11.13 ± 2.43)%) (t = 0.781-5.025, all P < 0.05).

CONCLUSIONThe sleeve gastrectomy procedure can improve the renal function in a diabetes rat model may be through protecting the podocytes function and preventing the mesangial expansion of glomeruli.

Animals ; Blood Glucose ; Creatinine ; blood ; urine ; Diabetes Mellitus, Experimental ; physiopathology ; Diabetic Nephropathies ; physiopathology ; surgery ; Gastrectomy ; Glomerular Filtration Rate ; Kidney ; physiopathology ; Kidney Function Tests ; Random Allocation ; Rats

Type 1 diabetes is an autoimmune disease caused by permanent destruction of insulin-producing pancreatic beta cells and requires lifelong exogenous insulin therapy. Recently, islet transplantation has been developed, and although there have been significant advances, this approach is not widely used clinically due to the poor survival rate of the engrafted islets. We hypothesized that improving survival of engrafted islets through ex vivo genetic engineering could be a novel strategy for successful islet transplantation. We transduced islets with adenoviruses expressing betacellulin, an epidermal growth factor receptor ligand, which promotes beta-cell growth and differentiation, and transplanted these islets under the renal capsule of streptozotocin-induced diabetic mice. Transplantation with betacellulin-transduced islets resulted in prolonged normoglycemia and improved glucose tolerance compared with those of control virus-transduced islets. In addition, increased microvascular density was evident in the implanted islets, concomitant with increased endothelial von Willebrand factor immunoreactivity. Finally, cultured islets transduced with betacellulin displayed increased proliferation, reduced apoptosis and enhanced glucose-stimulated insulin secretion in the presence of cytokines. These experiments suggest that transplantation with betacellulin-transduced islets extends islet survival and preserves functional islet mass, leading to a therapeutic benefit in type 1 diabetes.
Animals ; Apoptosis ; Betacellulin ; Cell Proliferation ; Diabetes Mellitus, Experimental/*surgery ; Glucose Intolerance/*surgery ; Humans ; Insulin-Secreting Cells/*metabolism/physiology ; Intercellular Signaling Peptides and Proteins/genetics/*metabolism ; *Islets of Langerhans Transplantation ; Mice ; Mice, Inbred C57BL ; Rats

OBJECTIVETo observe the influence of negative pressure wound therapy on the angiogenesis of wounds in diabetic rats.

METHODSDiabetes model was reproduced by intraperitoneal injection of 20 g/L streptozotocin in the dosage of 65 mg/kg in 40 SD rats. Two weeks later, rats were divided into control group (C) and negative pressure group (NP) according to the random number table, with 20 rats in each group. A piece of full-thickness skin in the center of the back of each rat in the size of 2 cm×2 cm was excised to produce a wound. Immediately after injury, wounds in group C were given conventional dressing change; wounds in group NP were treated with continuous negative pressure (-16.0 kPa) therapy for four hours a day, which lasted for seven days. (1) Blood glucose and body weight of rats in two groups were respectively measured by glucose meter and electronic scale before treatment, and 1 and 2 week (s) after. (2) Wound blood flow was detected by laser Doppler perfusion imager before treatment and on post treatment day (PTD) 1, 3, 7, with 5 rats at each time point. (3) On PTD 3 and 7, respectively, five rats from each group were sacrificed. The wound tissue was excised and divided into two parts. The angiogenesis in the left part tissue was observed with immunohistochemical staining. The microvessel density was calculated. (4) The full-thickness skin excised before treatment and the right part tissue freeze on PTD 3 and 7 were collected. On PTD 1 and 14, wound tissue was excised in the above-mentioned method. The mRNA levels of the vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 1 (Fit-1), angiopoietin 1 (Ang-1), Ang-2, and tyrosine kinase receptor 2 (Tie-2) were determined with real-time fluorescence quantification PCR. Data were processed with two-way analysis of variance or LSD-t test.

RESULTS(1) No significant difference was observed between two groups in blood glucose level and body weight as a whole or at each time point (with F values respectively 0.667, 0.176, t values from 0.311 to 0.707, P values all above 0.05). (2) The difference in the overall wound blood flow of rats between two groups was significant (F = 24.66, P < 0.05). On PTD 1, 3, 7, values of wound blood flow of rats in group NP were (179 ± 24), (219 ± 12), (192 ± 30) perfusion unit, significantly higher than those of rats in group C[(127 ± 16), (179 ± 8), (144 ± 17) perfusion unit, with t values respectively 3.71, 5.57, 2.77, P < 0.05 or P < 0.01]. (3) The difference in the overall microvessel density in the wound of rats between two groups was significant (F = 33.25, P < 0.05). On PTD 3, the microvessel density in the wound of rats in group NP was (80 ± 12) per 100-time visual field, which was significantly higher than that of group C[(38 ± 4) per 100-time visual field, t = 9.257, P < 0.05]. On PTD 7, the microvessel density in the wound of rats in two groups were close (t = 1.159, P > 0.05), but the vessels in group NP were regularly arranged with spacious lumen, while the vessels in group C were disorderly arranged with narrow lumen. (4) On PTD 1, 3, mRNA expression levels of VEGF, Fit-1, and Ang-1 in group NP were obviously higher than those in group C (with t values from 1.28 to 11.60, P values all below 0.01). On PTD 7, the mRNA expression level of Ang-1 (27.59 ± 3.55) in group NP was obviously higher than that in group C (19.87 ± 1.86, t = 7.23, P < 0.001), while the mRNA level of its antagonist Ang-2 (5.79 ± 0.61) in group NP was obviously lower than that in group C (17.62 ± 0.85, t = 19.88, P < 0.001). On PTD 3, 7, 14, mRNA levels of Tie-2 in group NP were obviously lower than those in group C (with t values from 8.92 to 15.60, P values all below 0.01).

CONCLUSIONSNegative pressure wound therapy may promote wound angiogenesis by enhancing the expression of Ang-1 and lowering the expression of Ang-2 in diabetic rats.

Angiopoietin-1 ; metabolism ; Angiopoietin-2 ; metabolism ; Animals ; Diabetes Mellitus, Experimental ; surgery ; Male ; Negative-Pressure Wound Therapy ; Neovascularization, Physiologic ; Rats ; Rats, Sprague-Dawley ; Wound Healing

OBJECTIVETo study the effect of pretreatment with apoptotic donor spleen cells on spleen lymphocyte function of recipient rats undergoing islet transplantation to explore new approaches to prolong islet graft survival.

METHODSApoptotic spleen cells from donor rats were obtained by exposure to γ-ray irradiation from (60)Co. Diabetic SD rat models were randomly divided into 4 groups to receive tail vein injections with saline (group A), normal cells (group B), apoptotic donor cells (group C), or necrotic donor cells (group D). One week later, orthotopic transplantation of islets under the renal capsule was performed. Before and at 1 and 2 weeks after islet transplantation, the recipient rats were examined for proliferative activity of spleen lymphocytes with CFSE cell staining and for IL-2 and IL-10 expressions in the cells using ELISA.

RESULTSPretreatment with donor apoptotic cells significantly suppressed the proliferative activity of recipient spleen lymphocytes before and at 1 and 2 weeks after islet transplantation as compared with the other three groups (P<0.05). The level of IL-2 was significantly decreased while IL-10 increased in apoptotic donor cell pretreatment group compared with those in the other 3 groups at each time point of observation.

CONCLUSIONThe effect of pretreatment with apoptotic donor cells on recipient spleen lymphocytes suggest an important role of apoptotic donor spleen cells in immune tolerance of grafts.

Animals ; Apoptosis ; immunology ; Cell Proliferation ; Cobalt Radioisotopes ; Diabetes Mellitus, Experimental ; metabolism ; pathology ; surgery ; Gamma Rays ; Graft Survival ; Immune Tolerance ; Interleukin-10 ; metabolism ; Interleukin-2 ; metabolism ; Islets of Langerhans Transplantation ; Lymphocyte Transfusion ; Lymphocytes ; metabolism ; pathology ; radiation effects ; Male ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Spleen ; cytology ; metabolism ; radiation effects

OBJECTIVETo investigate the relationship between the type 1 diabetic rats residual islet function and postoperative glycemia of gastric bypass procedure (GBP).

METHODSIntraperitoneal injection of STZ was used to produce type 1 diabetic rat model. According to the level of serum glucose, rats were divided into two groups: group 1 (fasting glucose 16.7-22.0 mmol/L, n=42) and group 2 (fasting glucose>22.0 mmol/L, n=54). Half rats of group 1 and group 2 received GBP, which were OP1 group (n=21) and OP2 group (n=27). The normal control group included 20 Wistar rats. The fasting glycemia and fasting C-peptide (C-P) were tested at postoperative weeks 1, 2, 3, and pancreas pathological slices were examined 3 weeks after surgery under microscope.

RESULTSAfter GBP, the C-P was elevated and the glycemia was well controlled in OP1 group compared with group 1 (P<0.05). But the C-P was not significantly increased and the glycemia control was poor compared with group 2 (P>0.05). Pathological examination revealed that there were partial islets residual in pancrease of group 1, the islets were shown obvious hyperplasia in OP1 group after GBP. There were almost no islets residual in pancrease of group 2, and the islets were shown no obvious hyperplasia in OP2 group after GBP.

CONCLUSIONSResidual islet function determines the glycemia changes of type 1 diabetic rats after gastric bypass.

Animals ; Blood Glucose ; metabolism ; Diabetes Mellitus, Experimental ; metabolism ; physiopathology ; surgery ; Female ; Gastric Bypass ; Male ; Pancreas ; physiopathology ; Postoperative Period ; Rats ; Rats, Wistar

OBJECTIVETo investigate the effects of gastric bypass surgery(GBP) on hepatic phosphoenolpyruvate carboxykinase(PEPCK) mRNA expression in type 2 diabetic Goto-Kakizaki rats.

METHODSMale GK rats were randomized into three groups: gastric bypass surgery(n=10), sham operation with diet restriction(n=10), and sham operation alone(n=10). Liver specimens of GK rats were collected during the intraoperative period for self-control study and 8 weeks after surgery. Fasting blood glucose, food intake, and body weight were recorded before surgery and 1, 2, 4, 8 weeks after surgery. The expression of PEPCK mRNA was measured by real-time PCR.

RESULTSThe fasting plasma glucose level decreased from(17.6±2.1) mmol/L before surgery to(7.5±0.9) mmol/L 8 weeks after surgery in GBP group. The level of PEPCK mRNA decreased from 1.08±0.38 before surgery to 0.41±0.10 8 weeks after surgery, significantly lower than that in sham operation alone group(1.04±0.12)(P<0.01). The level of PEPCK mRNA in diet restriction group increased from 1.15±0.16 before surgery to 2.54±0.82 8 weeks after surgery(P<0.01). The expression of PEPCK mRNA in diet restriction was significantly higher than that in CBP group(P<0.01).

CONCLUSIONSGBP can significantly improve hyperglycemia in type 2 diabetic GK rat models, which may be associated with the decrease of hepatic PEPCK mRNA level.

Animals ; Blood Glucose ; analysis ; Diabetes Mellitus, Experimental ; enzymology ; surgery ; Diabetes Mellitus, Type 2 ; enzymology ; surgery ; Gastric Bypass ; Intracellular Signaling Peptides and Proteins ; genetics ; metabolism ; Liver ; enzymology ; Male ; Phosphoenolpyruvate Carboxykinase (GTP) ; genetics ; metabolism ; RNA, Messenger ; genetics ; Rats

OBJECTIVETo investigate the influence and significance of gastric bypass surgery on hepatic gluconeogenesis in type 2 diabetic Goto Kakizaki(GK) rats.

METHODSForty GK rats were randomly divided into Roux-en-Y gastric bypass group(group A) and sham operation group(group B). Differences in glucose tolerance experiment(OGTT) at preoperative and postoperative 1, 2 and 4 weeks were compared and weight was recorded. Glycated hemoglobin levels were measured preoperatively and 4 weeks postoperatively. The animals were sacrificed 4 weeks after surgery and liver tissues were harvested to detect the relative expression of mRNA and protein of glucose 6 phosphatase(G-6-P) and phosphoenol pyruvate kinase(PEPCK) with RT-PCR and Western blot.

RESULTSFasting blood glucose levels were 6.5, 4.9, and 4.7 mmol/L in group A, and were 10.3, 10.4, and 12.5 mmol/L in group B, and the differences between two groups were statistically significant(P<0.05). The blood glucose level at 2 h after stomach lavage were 8.3, 6.4 and 5.5 mmol/L in group A, and were 21.4, 23.8 and 24.7 mmol/L in group B at postoperative 1, 2, 4 weeks, and the differences between two groups were statistically significant(P<0.05). The glycosylated hemoglobin at postoperative 4 weeks was(6.8±1.0)%, significantly lower than that in group B[(7.9±0.8)%, P<0.05]. Hepatic G-6-P and PEPCK mRNA relative expression at postoperative 4 weeks was reduced by 21.0% and 25.9% respectively as compared to group B, and the protein expression reduced as well. Immunohistochemistry showed that hepatic glycogen sedimentary in group A increased significantly.

CONCLUSIONThe relative mRNA and protein level of key enzymes of hepatic gluconeogenesis are significantly decreased after Roux-en-Y gastric bypass surgery and hepatic gluconeogenesis is reduced, which may be a potential mechanism of the decrease of blood glucose.

Animals ; Blood Glucose ; analysis ; Diabetes Mellitus, Experimental ; metabolism ; surgery ; Diabetes Mellitus, Type 2 ; metabolism ; surgery ; Gastric Bypass ; Gluconeogenesis ; Glucose-6-Phosphatase ; metabolism ; Glycated Hemoglobin A ; metabolism ; Intracellular Signaling Peptides and Proteins ; metabolism ; Liver ; enzymology ; Male ; Phosphoenolpyruvate Carboxykinase (GTP) ; metabolism ; Rats

Recent evidence has suggested that human skin fibroblasts may represent a novel source of therapeutic stem cells. In this study, we report a 3-stage method to induce the differentiation of skin fibroblasts into insulin-producing cells (IPCs). In stage 1, we establish the isolation, expansion and characterization of mesenchymal stem cells from human labia minora dermis-derived fibroblasts (hLMDFs) (stage 1: MSC expansion). hLMDFs express the typical mesenchymal stem cell marker proteins and can differentiate into adipocytes, osteoblasts, chondrocytes or muscle cells. In stage 2, DMEM/F12 serum-free medium with ITS mix (insulin, transferrin, and selenite) is used to induce differentiation of hLMDFs into endoderm-like cells, as determined by the expression of the endoderm markers Sox17, Foxa2, and PDX1 (stage 2: mesenchymal-endoderm transition). In stage 3, cells in the mesenchymal-endoderm transition stage are treated with nicotinamide in order to further differentiate into self-assembled, 3-dimensional islet cell-like clusters that express multiple genes related to pancreatic beta-cell development and function (stage 3: IPC). We also found that the transplantation of IPCs can normalize blood glucose levels and rescue glucose homeostasis in streptozotocin-induced diabetic mice. These results indicate that hLMDFs have the capacity to differentiate into functionally competent IPCs and represent a potential cell-based treatment for diabetes mellitus.
Animals ; Biological Markers/metabolism ; *Cell Culture Techniques ; *Cell Differentiation ; Cell Proliferation/drug effects ; Cell Separation ; Cells, Cultured ; Dermis/*cytology/drug effects ; Diabetes Mellitus, Experimental/*surgery ; Female ; Fibroblasts/*cytology/drug effects ; Genitalia, Female/*cytology ; Glucose/metabolism ; Hepatocyte Nuclear Factor 3-beta/metabolism ; Homeodomain Proteins/metabolism ; Humans ; Insulin/pharmacology/secretion ; Insulin-Secreting Cells/*cytology/metabolism ; *Islets of Langerhans Transplantation ; Mesenchymal Stem Cells/*cytology/drug effects/metabolism ; Mice ; Mice, Nude ; Niacinamide/pharmacology ; Recovery of Function ; SOXF Transcription Factors/metabolism ; Sodium Selenite/pharmacology ; Trans-Activators/metabolism ; Transferrin/pharmacology

OBJECTIVETo study the protective effect of islet xenograft and its possible mechanism of high expression of heme oxygenase-1 (HO-1) in donor pancreas islet induced by cobalt protoporphyrin (CoPP).

METHODSMale SD rats and C57BL/6 mouse were used as donors and recipients respectively. Donors were divided into 3 groups according to different pretreatment 24 hours before donation: control group (injected intraperitoneally with NaCl), induce group [injected intraperitoneally with cobalt-protoporphyrin (CoPP)], block group (injected intraperitoneally with CoPP and zinc protoporphyrin simultaneously). A modified approach was used for islet isolation.Recipients were rendered diabetic by intraperitoneal injection of streptozotocin. Islets were transplanted into mouse subrenal capsule. Postoperative mouse glycemia were monitored daily and normoglycemia time was compared among each group. The receptor mouse serum IL-10 was detected by ELISA approach, and real-time PCR was used to check the expression of IL-10 mRNA in islet graft tissues. The graft tissues were observed for the lymphocyte infiltration after HE staining.

RESULTSDiabetes mice accepted islets untreated, induced or blocked maintained the euglycemia for (9.3 +/- 1.4), (16.3 +/- 1.5) and (9.7 +/- 1.0) d respectively. The xeno-islets presented HO-1 over-expression survived much longer than that absent (P < 0.05), it was no significance between control group and block group (P > 0.05). The mouse islet serum IL-10 content after induction was (73.0 +/- 9.7) pg/ml, significantly higher than (30.6 +/- 3.9) pg/ml of the untreated group and (32.1 +/- 5.9) pg/ml of the blocked group (P < 0.05), there was no difference between control group and block group (P > 0.05). Moreover, the IL-10 mRNA expression up-regulated statistic significantly in HO-1 induced islet xeno-graft. Pathological examination showed that the graft lymphocyte infiltration of the induced group was obviously less serious than the other two groups.

CONCLUSIONSThe higher expression of HO-1 induced by CoPP in vivo would significantly prolong graft survival time and its mechanism could be related to immune modulation of IL-10.

Animals ; Diabetes Mellitus, Experimental ; metabolism ; pathology ; surgery ; Graft Survival ; Heme Oxygenase-1 ; drug effects ; metabolism ; Interleukin-10 ; metabolism ; Islets of Langerhans ; metabolism ; pathology ; Male ; Mice ; Mice, Inbred C57BL ; Pancreas Transplantation ; Protoporphyrins ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Subrenal Capsule Assay ; Transplantation, Heterologous

OBJECTIVETo establish a new gastric bypass animal-model with Goto-Kakizaki rats whose different parts of the small intestine were bypassed while stomach was not bypassed.

METHODSForty male 3-month-old GK rats were randomly divided into 4 groups: group I (sham operation), group II (duodenum bypassed), group III (jejunum bypassed), group IV (ileum bypassed). Fasting plasma glucose was measured before operation and the 1st, 4th,and 8th week after operation in all the rats, the body weight of all the rats were measured simultaneously.

RESULTSThe survival rate of operation for the rats was 95%. Two rats in group IV (died on the first day after operation. The mean fasting plasma glucose concentration of the rats in group II, III, IV (declined obviously 4 weeks after gastric bypass [group II (12.02+/-1.97) vs (6.36+/-0.50) mmol/L, group III (13.42+/-1.66) vs (5.96+/-0.53) mmol/L, group IV (14.32+/-2.82) vs (5.18+/-0.49) mmol/L, all P <0.01], but there were no significant differences among the gastric bypassed groups. The weight of rats in group I, II, III (increased obviously after gastric bypass [group I (253.6+/-9.37) vs (367.0+/-23.70) g, group II (268.2+/-7.95) vs (384.8+/-16.12) g, group III (253.0+/-6.20) vs (323.0+/-16.40) g, all P <0.05] except the rats in group IV ([(262.0+/-13.47) vs(185.8+/-11.56) g].

CONCLUSIONSThe mean fasting plasma glucose concentration of the GK rats decreases obviously after gastric bypass through different parts of small intestine. The fasting plasma glucose concentration is not associated with the length of small intestine and body weight.

Animals ; Blood Glucose ; analysis ; Diabetes Mellitus, Experimental ; surgery ; Diabetes Mellitus, Type 2 ; surgery ; Gastric Bypass ; Male ; Models, Animal ; Rats ; Rats, Inbred Strains