Effect of Sinapic Acid on Scopolamine-Induced Learning and Memory Impairment in SD Rats.

The seriousness of the diseases caused by aging have recently gained attention. Alzheimer's disease (AD), a chronic neurodegenerative disease, accounts for 60-80% of senile dementia cases. Continuous research is being conducted on the cause of Alzheimer's disease, and it is believed to include complex factors, such as genetic factors, the accumulation of amyloid beta plaques, a tangle of tau protein, oxidative stress, cholinergic dysfunction, neuroinflammation, and cell death. Sinapic acid is a hydroxycinnamic acid found in plant families, such as oranges, grapefruit, cranberry, mustard seeds, and rapeseeds. It exhibits various biological activities, including anti-inflammatory, anti-oxidant, anti-cancer, and anti-depressant effects. Sinapic acid is an acetylcholine esterase inhibitor that can be applied to the treatment of dementia caused by Alzheimer's disease and Parkinson's disease. However, electrophysiological studies on the effects of sinapic acid on memory and learning must still be conducted. Therefore, it was confirmed that sinapic acid was effective in long-term potentiation (LTP) using organotypic hippocampal segment tissue. In addition, the effect on scopolamine-induced learning and memory impairment was measured by oral administration of sinapic acid 10 mg/kg/day for 14 days, and behavioral experiments related to short-term and long-term spatial memory and avoidance memory were conducted. Sinapic acid increased the activity of the field excitatory postsynaptic potential (fEPSP) in a dose-dependent manner after TBS, and restored fEPSP activity in the CA1 region suppressed by scopolamine. The scopolamine-induced learning and memory impairment group showed lower results than the control group in the Y-maze, Passive avoidance (PA), and Morris water maze (MWM) experiments. Sinapic acid improved avoidance memory, short and long-term spatial recognition learning, and memory. In addition, sinapic acid weakened the inhibition of the brain-derived neurotrophic factor (BDNF), tropomyosin receptor kinase B (TrkB) and the activation of prostaglandin-endoperoxide synthase 2 (COX-2) and interleukin 1 beta (IL-1ß) induced by scopolamine in the hippocampus. These results show that sinapic acid is effective in restoring LTP and cognitive impairment induced by the cholinergic receptor blockade. Moreover, it showed the effect of alleviating the reduction in scopolamine-induced BDNF and TrkB, and alleviated neuroinflammatory effects by inhibiting the increase in COX-2 and IL-1ß. Therefore, we showed that sinapic acid has potential as a treatment for neurodegenerative cognitive impairment.

Comparison of Outcomes of Multi-Level Anterior, Oblique, Transforaminal Lumbar Interbody Fusion Surgery : Impact on Global Sagittal Alignment.

OBJECTIVE: To compare the outcomes of anterior lumbar interbody fusion (ALIF), oblique lumbar interbody fusion (OLIF), and transforaminal lumbar interbody fusion (TLIF) in terms of global sagittal alignment. METHODS: From January 2007 to December 2019, 141 adult patients who underwent multilevel interbody fusion for lumbar degenerative disorders were enrolled. Regarding the approach, patients were divided into the ALIF (n=23), OLIF (n=60), and TLIF (n=58) groups. Outcomes, including local radiographic parameters and global sagittal alignment, were then compared between the treatment groups. RESULTS: Regarding local radiographic parameters, ALIF and OLIF were superior to TLIF in terms of the change in the anterior disc height (7.6±4.5 mm vs. 6.9±3.2 mm vs. 4.7±2.9 mm, p<0.001), disc angle (-10.0°±6.3° vs. -9.2°±5.2° vs. -5.1°±5.1°, p<0.001), and fused segment lordosis (-14.5°±11.3° vs. -13.8°±7.5° vs. -7.4°±9.1°, p<0.001). However, regarding global sagittal alignment, postoperative lumbar lordosis (-42.5°±9.6° vs. -44.4°±11.6° vs. -40.6°±12.3°, p=0.210), pelvic incidence-lumbar lordosis mismatch (7.9°±11.3° vs. 6.7°±11.6° vs. 11.5°±13.0°, p=0.089), and the sagittal vertical axis (24.3±28.5 mm vs. 24.5±34.0 mm vs. 25.2±36.6 mm, p=0.990) did not differ between the groups. CONCLUSION: Although the anterior approaches were superior in terms of local radiographic parameters, TLIF achieved adequate global sagittal alignment, comparable to the anterior approaches.

Photoswitchable Microgels for Dynamic Macrophage Modulation.

Dynamic manipulation of supramolecular self-assembled structures is achieved irreversibly or under non-physiological conditions, thereby limiting their biomedical, environmental, and catalysis applicability. In this study, microgels composed of azobenzene derivatives stacked via π-cation and π-π interactions are developed that are electrostatically stabilized with Arg-Gly-Asp (RGD)-bearing anionic polymers. Lateral swelling of RGD-bearing microgels occurs via cis-azobenzene formation mediated by near-infrared-light-upconverted ultraviolet light, which disrupts intermolecular interactions on the visible-light-absorbing upconversion-nanoparticle-coated materials. Real-time imaging and molecular dynamics simulations demonstrate the deswelling of RGD-bearing microgels via visible-light-mediated trans-azobenzene formation. Near-infrared light can induce in situ swelling of RGD-bearing microgels to increase RGD availability and trigger release of loaded interleukin-4, which facilitates the adhesion structure assembly linked with pro-regenerative polarization of host macrophages. In contrast, visible light can induce deswelling of RGD-bearing microgels to decrease RGD availability that suppresses macrophage adhesion that yields pro-inflammatory polarization. These microgels exhibit high stability and non-toxicity. Versatile use of ligands and protein delivery can offer cytocompatible and photoswitchable manipulability of diverse host cells.

Effect of Dexmedetomidine Compared to Remifentanil During Bariatric Surgery on Postoperative Nausea and Vomiting: a Retrospective Study.

PURPOSE: Postoperative nausea and vomiting (PONV) occurs frequently after bariatric surgery and is a major cause of adverse outcomes. This retrospective study investigated whether opioid-restricted total intravenous anesthesia using dexmedetomidine as a substitute for remifentanil can reduce PONV in bariatric surgery. MATERIALS AND METHODS: The electronic medical records of adult patients who underwent laparoscopic bariatric surgery between January and December 2019 were reviewed. The patients were divided into two groups according to the agents used for anesthesia: Group D, propofol and dexmedetomidine; Group R, propofol and remifentanil. RESULTS: A total of 134 patients were included in the analyses. The frequency of postoperative nausea was significantly lower in Group D than that in Group R until 2 h after discharge from the postanesthesia care unit (PACU) (P = 0.005 in the PACU, P = 0.010 at 2 h after PACU discharge) but failed to significantly reduce the overall high incidence rates of 60.5% and 65.5%, respectively (P = 0.592). Postoperative pain score was significantly lower in Group D until 6 h after PACU discharge. The rates of rescue antiemetic and analgesic agent administration in the PACU were significantly lower in Group D than those in Group R. CONCLUSION: Opioid-restricted total intravenous anesthesia using dexmedetomidine reduces postoperative nausea, pain score, antiemetic, and analgesic requirements in the immediate postoperative period after bariatric surgery.

5-HT3 receptor antagonists decrease the prevalence of postoperative delirium in older patients undergoing orthopedic lower limb surgery.

BACKGROUND: Delirium is an important postoperative complication. Recent research suggested that 5-hydroxytryptamine 3 (5-HT3) receptor antagonists may have clinical effect in the treatment and prevention of delirium. We investigated the association between 5-HT3 receptor antagonists and the occurrence of postoperative delirium (POD). METHODS: Retrospectively, the electronic medical records were reviewed in patients aged ≥ 65 years who underwent orthopedic lower limb surgery under regional anesthesia (spinal or combined spinal-epidural anesthesia) and administered intravenous 0.075 mg palonosetron or 0.3 mg ramosetron prior to the end of surgery between July 2012 and September 2015. POD incidence and anesthesia-, surgery-, and patient-related factors were evaluated. To investigate the association between 5-HT3 receptor antagonists and the occurrence of POD, multivariable logistic regression analysis was performed. RESULTS: Of the 855 patients included, 710 (83%) were administered 5-HT3 receptor antagonists. POD was confirmed in 46 (5.4%) patients. 5-HT3 receptor antagonists reduced the POD incidence by 63% (odds ratio [OR] 0.37; 95% confidence interval [CI], 0.15-0.94; P = 0.04). Moreover, the POD incidence decreased by 72% (OR 0.28, 95% CI 0.10-0.77, P = 0.01) when palonosetron was administered. Other identified risk factors for POD were emergency surgery, older age, hip surgery, lower body mass index, and intraoperative propofol sedation. CONCLUSION: 5-HT3 receptor antagonists may be related with a significantly reduced risk for POD in older patients undergoing orthopedic lower limb surgery. Notably, palonosetron was more effective for POD prevention.

Skimmed milk fermented by lactic acid bacteria inhibits adipogenesis in 3T3-L1 pre-adipocytes by downregulating PPARγ via TNF-α induction in vitro.

The murine 3T3-L1 pre-adipocyte cell line is widely used as an in vitro model for adipogenesis because of its similarities to primary fat cells. The aim of this study was to investigate the intracellular mechanisms by which skimmed milk fermented by two lactic acid bacteria (LAB), Enterococcus faecalis and Lactobacillus plantarum, inhibited the differentiation of 3T3-L1 pre-adipocytes. Skimmed milk fermented by both LAB, but not non-fermented skimmed milk, significantly reduced the accumulation of lipid droplets and cellular triglycerides in a concentration-dependent manner. The mRNA and protein levels of peroxisome proliferator-activated receptor γ (PPARγ) were markedly inhibited in the presence of skimmed milk fermented by both LAB. Furthermore, the skimmed milk fermented by both LAB decreased the mRNA and protein expressions of PPARγ-targeting genes, lipoprotein lipase and adipocyte fatty acid-binding protein. Under the same circumstances, resistin mRNA expression was downregulated, but not leptin mRNA expression. In contrast, skimmed milk fermented by both LAB significantly upregulated tumor necrosis factor-α (TNF-α). These results suggest that LAB-fermented skimmed milk inhibits adipogenesis by inhibiting a master transcription factor PPARγ via the upregulation of the proinflammatory cytokine TNF-α in 3T3-L1 cells.

Common causes and characteristics of adverse drug reactions in older adults: a retrospective study.

BACKGROUND: Aging populations are often accompanied by comorbidity and polypharmacy, leading to increases in adverse drug reactions (ADRs). We sought to evaluate the causes and characteristics of ADRs in older Korean adults (≥65 years) in comparison to younger individuals (< 65 years). METHODS: Of 37,523 cases reported at a Korean pharmacovigilance center from 2011 to 2018, we reviewed 18,842 ADRs of certain or probable causality on the basis of WHO-UMC criteria. We estimated the number of ADRs per 1000 patients exposed to the major culprit drugs, and incidence rate ratios were obtained to assess high- and low-risk medications in older adults. RESULTS: In total, 4152 (22.0%) ADRs were reported for 3437 older adults (mean age, 74.6 years and 57.3% female). Tramadol (rate ratio, 1.32; 95% confidence interval [CI], 1.21-1.44; P < 0.001) and fentanyl (1.49, 1.16-1.92, P = 0.002) posed higher risks of ADRs in the older adults, whereas nonsteroidal anti-inflammatory drugs (NSAIDs) (0.35, 0.30-0.40, P < 0.001) and iodinated contrast media (ICM) (0.82, 0.76-0.89, P < 0.001) posed lower risks. Ratios of serious ADRs to NSAIDs (odds ratio, 2.16; 95% CI, 1.48-3.15; P < 0.001) and ICM (2.09, 1.36-3.21, P = 0.001) were higher in the older adults than in the younger patients. Analgesics primarily elicited cutaneous ADRs in the younger patients and gastrointestinal reactions in the older adults. ICM more commonly led to anaphylaxis in the older adults than the younger patients (3.0% vs. 1.6%, P = 0.019). CONCLUSION: For early detection of ADRs in older adults, better understanding of differences in the causes and characteristics thereof in comparison to the general population is needed.

A Prospective Observation of Psychological Distress in Patients With Anaphylaxis.

PURPOSE: Anaphylaxis is an immediate allergic reaction characterized by potentially life-threatening, severe, systemic manifestations. While studies have evaluated links between serious illness and posttraumatic stress disorder (PTSD), few have investigated PTSD after anaphylaxis in adults. We sought to investigate the psychosocial burden of recent anaphylaxis in Korean adults. METHODS: A total of 203 (mean age of 44 years, 120 females) patients with anaphylaxis were recruited from 15 university hospitals in Korea. Questionnaires, including the Impact of Event Scale-Revised-Korean version (IES-R-K), the Korean version of the Beck Anxiety Inventory (K-BAI), and the Korean version of the Beck Depression Inventory (K-BDI), were administered. Demographic characteristics, causes and clinical features of anaphylaxis, and serum inflammatory markers, including tryptase, platelet-activating factor, interleukin-6, tumor necrosis factor-α, and C-reactive protein, were evaluated. RESULTS: PTSD (IES-R-K ≥ 25) was noted in 84 (41.4%) patients with anaphylaxis. Of them, 56.0% had severe PTSD (IES-R-K ≥ 40). Additionally, 23.2% and 28.1% of the patients had anxiety (K-BAI ≥ 22) and depression (K-BDI ≥ 17), respectively. IES-R-K was significantly correlated with both K-BAI (r = 0.609, p < 0.0001) and K-BDI (r = 0.550, p < 0.0001). Among the inflammatory mediators, tryptase levels were lower in patients exhibiting PTSD; meanwhile, platelet-activating factor levels were lower in patients exhibiting anxiety and depression while recovering from anaphylaxis. In multivariate analysis, K-BAI and K-BDI were identified as major predictive variables of PTSD in patients with anaphylaxis. CONCLUSIONS: In patients with anaphylaxis, we found a remarkably high prevalence of PTSD and associated psychological distresses, including anxiety and depression. Physicians ought to be aware of the potential for psychological distress in anaphylactic patients and to consider psychological evaluation.

Enhancement of mouse sperm motility by trophinin-binding peptide.

BACKGROUND: Trophinin is an intrinsic membrane protein that forms a complex in the cytoplasm with bystin and tastin, linking it microtubule-associated motor dynein (ATPase) in some cell types. Previously, we found that human sperm tails contain trophinin, bystin and tastin proteins, and that trophinin-binding GWRQ (glycine, tryptophan, arginine, glutamine) peptide enhanced motility of human sperm. METHODS: Immunohistochemistry was employed to determine trophinin protein in mouse spermatozoa from wild type mouse, by using spermatozoa from trophinin null mutant mice as a negative control. Multivalent 8-branched GWRQ (glycine, tryptophan, arginine, glutamine) peptide or GWRQ-MAPS, was chemically synthesized, purified by HPLC and its structure was confirmed by MALDI-TOF mass spectrometry. Effect of GWRQ-MAPS on mouse spermatozoa from wild type and trophinin null mutant was assessed by a computer-assisted semen analyzer (CASA). RESULTS: Anti-trophinin antibody stained the principal (central) piece of the tail of wild type mouse sperm, whereas the antibody showed no staining on trophinin null sperm. Phage particles displaying GWRQ bound to the principal piece of sperm tail from wild type but not trophinin null mice. GWRQ-MAPS enhanced motility of spermatozoa from wild type but not trophinin null mice. CASA showed that GWRQ-MAPS enhanced both progressive motility and rapid motility in wild type mouse sperm. CONCLUSIONS: Present study established the expression of trophinin in the mouse sperm tail and trophinin-dependent effect of GWRQ-MAPS on sperm motility. GWRQ causes a significant increase in sperm motility.

Remission of diabetes by insulin gene therapy using a hepatocyte-specific and glucose-responsive synthetic promoter.

Efficient production of insulin in response to changes in glucose levels has been a major issue for insulin gene therapy to treat diabetes. To express target genes in response to glucose specifically in hepatocytes, we generated a synthetic promoter library containing hepatocyte nuclear factor-1, CAAT/enhancer-binding protein (C/EBP) response element, and glucose-response element. Combinations of these three cis-elements in 3-, 6-, or 9-element configurations were screened for transcriptional activity and then glucose responsiveness in vitro. The most effective promoter (SP23137) was selected for further study. Intravenous administration of a recombinant adenovirus expressing furin-cleavable rat insulin under control of the SP23137 promoter into streptozotocin (STZ)-induced diabetic mice resulted in normoglycemia, which was maintained for >30 days. Glucose tolerance tests showed that treated mice produced insulin in response to glucose and cleared exogenous glucose from the blood in a manner similar to nondiabetic control mice, although the clearance was somewhat delayed. Insulin expression was seen specifically in the liver and not in other organs. These observations indicate the potential of this synthetic, artificial promoter to regulate glucose-responsive insulin production and remit hyperglycemia, thus providing a new method of liver-directed insulin gene therapy for type 1 diabetes.

Remission of diabetes by beta-cell regeneration in diabetic mice treated with a recombinant adenovirus expressing betacellulin.

Type 1 diabetes results from destruction of the majority of the pancreatic beta cells by beta cell-specific autoimmune responses; therefore, expansion of the beta-cell mass in vivo is a possible approach to its treatment. Betacellulin (BTC) is known to promote beta-cell growth and differentiation. We investigated whether transient, constitutive expression, and secretion of BTC would regenerate sufficient numbers of pancreatic beta cells to restore normoglycemia in diabetic animals. We constructed a recombinant adenoviral vector (rAd-BTC) containing the cytomegalovirus promoter/enhancer, beta-globin chimeric intron, and albumin leader sequence to facilitate secretion, followed by BTC (1-80) complementary DNA (cDNA) encoding mature BTC. A single intravenous (i.v.) administration of rAd-BTC resulted in complete remission of streptozotocin (STZ)-induced diabetes within 2 weeks in mice. The mice remained normoglycemic for >100 days; glucose tolerance tests showed kinetics similar to normal, nondiabetic mice. Pancreatic insulin content, beta-cell mass, and serum insulin levels in rAd-BTC-treated mice were significantly higher than in the controls. Treatment of autoimmune diabetic mice with rAd-BTC in combination with an immune suppressor resulted in remission of diabetes. We conclude that transient expression of BTC by rAd-BTC administration results in prolonged remission of diabetes in mice, by the regeneration of sufficient numbers of beta cells in the pancreas.

Remission of Diabetes by ß-Cell Regeneration in Diabetic Mice Treated With a Recombinant Adenovirus Expressing Betacellulin.

Type 1 diabetes results from destruction of the majority of the pancreatic ß cells by ß cell-specific autoimmune responses; therefore, expansion of the ß-cell mass in vivo Is a possible approach to its treatment. Betacellulin (BTC) is known to promote ß-cell growth and differentiation. We investigated whether transient, constitutive expression, and secretion of BTC would regenerate sufficient numbers of pancreatic ß cells to restore normoglycemia in diabetic animals. We constructed a recombinant adenoviral vector (rAd-BTC) containing the cytomegalovirus promoter/enhancer, ß-globin chimeric intron, and albumin leader sequence to facilitate secretion, followed by BTC (1-80) complementary DNA (cDNA) encoding mature BTC. A single intravenous (IV) administration of rAd-BTC resulted in complete remission of streptozotocin (STZ)-induced diabetes within 2 weeks in mice. The mice remained normoglycemic for >100 days; glucose tolerance tests showed kinetics similar to normal, nondiabetic mice. Pancreatic insulin content, ß-cell mass, and serum insulin levels in rAd-BTC-treated mice were significantly higher than in the controls. Treatment of autoimmune diabetic mice with rAd-BTC in combination with an immune suppressor resulted in remission of diabetes. We conclude that transient expression of BTC by rAd-BTC administration results in prolonged remission of diabetes in mice, by the regeneration of sufficient numbers of ß cells in the pancreas.

Human chorionic gonadotropin prevents Sjögren's syndrome-like exocrinopathy in mice.

OBJECTIVE: Results of recent studies suggest that human chorionic gonadotropin (HCG), a placental glycoprotein hormone required for the maintenance of pregnancy, may have immunomodulatory properties. Primary Sjögren's syndrome (SS), a chronic autoimmune disorder of unknown etiology, affects multiple exocrine glands including the salivary and lacrimal glands. The purpose of the present study was to determine whether HCG could prevent the development of salivary gland exocrinopathy in NOD mice, an experimental model of Sjögren's-like syndrome. METHODS: Female NOD mice were treated with HCG from 6 weeks of age to 12 weeks of age. At 14 weeks, tissue samples were evaluated for inflammatory lesions and cytokine messenger RNA by real-time polymerase chain reaction. At 18 weeks, the salivary flow rate was measured. RESULTS: Treatment with HCG resulted in a significant decrease in lymphocyte infiltration and parenchymal cell damage in the submandibular salivary glands. Messenger RNA levels of interferon-gamma, tumor necrosis factor alpha, interleukin-1beta, and interleukin-10, as well as inducible nitric oxide synthase and matrix metalloproteinase 9, were significantly decreased. Function studies revealed a marked increase in the salivary flow rate in HCG-treated mice compared with that in phosphate buffered saline-treated mice. CONCLUSION: In NOD mice, HCG acts as an immune modulator and prevents the development of salivary gland exocrinopathy. These findings suggest that HCG, a naturally occurring reproductive hormone, may be useful in the treatment of Sjögren's syndrome and other human autoimmune diseases.

Mechanisms involved in Korean mistletoe lectin-induced apoptosis of cancer cells.

AIM: To investigate the anti-cancer mechanisms of Korean mistletoe lectin (Viscum album coloratum agglutinin, VCA) using a human colon cancer cell line (COLO). METHODS: Cytotoxic effects of VCA on COLO cells were determined by 3- (4, 5-dimethylthiazol-2-yl) -2, 5-diphenyltetrazolium bromide (MTT) assay in vitro and tumor-killing effects in vivo. To study the mechanisms involved, the expression of various pro-caspases, anti-apoptotic proteins, and death receptors was determined by western blot. To determine which death receptor is involved in VCA-induced apoptosis of COLO cells, cytotoxicity was examined by MTT assay after treatment with agonists or antagonists of death receptors. RESULTS: VCA killed COLO cells in a time- and dose-dependent manner and induced complete regression of tumors in nude mice transplanted with COLO cells. Treatment of COLO cells with VCA activated caspase-2, -3, -8, and -9 and decreased expression of anti-apoptotic molecules including receptor interacting protein, nuclear factor-kappaB, X-linked inhibitor of apoptosis protein, and Akt/protein kinase B. We then examined the involvement of death receptors in VCA-induced apoptosis. Only tumor necrosis factor receptor 1, among the death receptors examined, was involved in apoptosis of COLO cells, evidenced by inhibition of VCA-induced apoptosis and decreased activation of caspases, particularly caspase-8, by tumor necrosis factor receptor 1 antagonizing antibody. CONCLUSION: VCA-induced apoptotic COLO cell death is due to the activation of caspases and inhibition of anti-apoptotic proteins, in part through the tumor necrosis factor receptor 1 signaling pathway.

Glucagon-like peptide-1 gene therapy in obese diabetic mice results in long-term cure of diabetes by improving insulin sensitivity and reducing hepatic gluconeogenesis.

Long-term treatment with glucagon-like peptide (GLP)-1 or its analog can improve insulin sensitivity. However, continuous administration is required due to its short half-life. We hypothesized that continuous production of therapeutic levels of GLP-1 in vivo by a gene therapy strategy may remit hyperglycemia and maintain prolonged normoglycemia. We produced a recombinant adenovirus expressing GLP-1 (rAd-GLP-1) under the cytomegalovirus promoter, intravenously injected it into diabetic ob/ob mice, and investigated the effect of this treatment on remission of diabetes, as well as the mechanisms involved. rAd-GLP-1-treated diabetic ob/ob mice became normoglycemic 4 days after treatment, remained normoglycemic over 60 days, and had reduced body weight gain. Glucose tolerance tests found that exogenous glucose was cleared normally. rAd-GLP-1-treated diabetic ob/ob mice showed improved beta-cell function, evidenced by glucose-responsive insulin release, and increased insulin sensitivity, evidenced by improved insulin tolerance and increased insulin-stimulated glucose uptake in adipocytes. rAd-GLP-1 treatment increased basal levels of insulin receptor substrate (IRS)-1 in the liver and activation of IRS-1 and protein kinase C by insulin in liver and muscle; increased Akt activation was only observed in muscle. rAd-GLP-1 treatment reduced hepatic glucose production and hepatic expression of phosphoenolpyruvate carboxykinase, glucose-6-phosphatase, and fatty acid synthase in ob/ob mice. Taken together, these results show that a single administration of rAd-GLP-1 results in the long-term remission of diabetes in ob/ob mice by improving insulin sensitivity through restoration of insulin signaling and reducing hepatic gluconeogenesis.

Engineered enteroendocrine cells secrete insulin in response to glucose and reverse hyperglycemia in diabetic mice.

Type 1 diabetes is a metabolic disorder caused by loss of insulin-producing pancreatic beta-cells. Expression of insulin in non-beta-cells to create beta-cell surrogates has been tried to treat type 1 diabetes. Enteroendocrine K cells have characteristics similar to pancreatic beta-cells, such as a glucose-sensing system and insulin-processing proteases. In this study, we genetically engineered an enteroendocrine cell line (STC-1) to express insulin under the control of the glucose-dependent insulinotropic polypeptide promoter. We screened clones and chose one, Gi-INS-7, based on its high production of insulin. Gi-INS-7 cells expressed glucose transporter 2 (GLUT2) and glucokinase (GK) and secreted insulin in response to elevated glucose levels in vitro. To determine whether Gi-INS-7 cells can control blood glucose levels in diabetic mice, we transplanted these cells under the kidney capsule of streptozotocin (STZ)-induced diabetic mice and found that blood glucose levels became normal within 2 weeks of transplantation. In addition, glucose tolerance tests in mice that became normoglycemic after transplantation with Gi-INS-7 cells showed that exogenous glucose was cleared appropriately. These results suggest that engineered K cells may be promising surrogate beta-cells for possible therapeutic use for the treatment of type 1 diabetes.

Prolonged remission of diabetes by regeneration of beta cells in diabetic mice treated with recombinant adenoviral vector expressing glucagon-like peptide-1.

Type 1 diabetes results from insulin deficiency caused by destruction of pancreatic beta cells. Glucagon-like peptide (GLP)-1 stimulates beta cell growth and differentiation. To determine whether continuous expression of GLP-1 in vivo can regenerate beta cells and remit type 1 diabetes in mice for a prolonged time, we constructed an adenoviral vector containing the cytomegalovirus promoter/enhancer and albumin leader sequence followed by GLP-1 cDNA (rAd-GLP-1). A single administration of rAd-GLP-1 via the tail vein into streptozotocin (STZ)-induced diabetic non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice resulted in remission of diabetes within 10 days; normoglycemia remained until the experiment was terminated. The number of insulin-positive cells in the pancreas and insulin secretion significantly increased in rAd-GLP-1-treated mice compared with STZ-induced diabetic mice treated with rAd-beta-galactosidase. Glucose tolerance tests in mice that achieved normoglycemia after treatment with rAd-GLP-1 showed that the kinetics of glucose clearance was similar to normal NOD/SCID mice. Treatment of autoimmune diabetic mice with rAd-GLP-1 restored normoglycemia, which was maintained for 1 year when mice were also treated with an immunoregulator to halt the autoimmune response to beta cells. We suggest that regeneration of insulin-producing cells by GLP-1 gene therapy may be a potential method for prolonged control of type 1 diabetes in humans.

Prolonged Remission of Diabetes by Regeneration of ß Cells in Diabetic Mice Treated with Recombinant Adenoviral Vector Expressing Glucagon-like Peptide-1.

Type 1 diabetes results from insulin deficiency caused by destruction of pancreatic ß cells. Glucagon-like peptide (GLP)-1 stimulates ß cell growth and differentiation. To determine whether continuous expression of GLP-1 in vivo can regenerate ß cells and remit type 1 diabetes in mice for a prolonged time, we constructed an adenoviral vector containing the cytomegalovirus promoter/enhancer and albumin leader sequence followed by GLP-1 cDNA (rAd-GLP-1). A single administration of rAd-GLP-1 via the tail vein into streptozotocin (STZ)-induced diabetic non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice resulted in remission of diabetes within 10 days; normoglycemia remained until the experiment was terminated. The number of insulin-positive cells in the pancreas and insulin secretion significantly increased in rAd-GLP-1-treated mice compared with STZ-induced diabetic mice treated with rAd-ß-galactosidase. Glucose tolerance tests in mice that achieved normoglycemia after treatment with rAd-GLP-1 showed that the kinetics of glucose clearance was similar to normal NOD/SCID mice. Treatment of autoimmune diabetic mice with rAd-GLP-1 restored normoglycemia, which was maintained for 1 year when mice were also treated with an immunoregulator to halt the autoimmune response to ß cells. We suggest that regeneration of insulin-producing cells by GLP-1 gene therapy may be a potential method for prolonged control of type 1 diabetes in humans.

Reversal of mouse hepatic failure using an implanted liver-assist device containing ES cell-derived hepatocytes.

Severe acute liver failure, even when transient, must be treated by transplantation and lifelong immune suppression. Treatment could be improved by bioartificial liver (BAL) support, but this approach is hindered by a shortage of human hepatocytes. To generate an alternative source of cells for BAL support, we differentiated mouse embryonic stem (ES) cells into hepatocytes by coculture with a combination of human liver nonparenchymal cell lines and fibroblast growth factor-2, human activin-A and hepatocyte growth factor. Functional hepatocytes were isolated using albumin promoter-based cell sorting. ES cell-derived hepatocytes expressed liver-specific genes, secreted albumin and metabolized ammonia, lidocaine and diazepam. Treatment of 90% hepatectomized mice with a subcutaneously implanted BAL seeded with ES cell-derived hepatocytes or primary hepatocytes improved liver function and prolonged survival, whereas treatment with a BAL seeded with control cells did not. After functioning in the BAL, ES cell-derived hepatocytes developed characteristics nearly identical to those of primary hepatocytes.

Viruses cause type 1 diabetes in animals.

More than 10 viruses have been reported to be associated with the development of type 1 diabetes-like symptoms in animals, with the best evidence coming from studies on the D variant of encephalomyocarditis (EMC-D) virus in mice and Kilham rat virus (KRV) in rats. A high titer of EMC-D viral infection results in the development of diabetes within 3 days, primarily due to the rapid destruction of beta cells by viral replication within the cells. A low titer of EMC-D viral infection results in the recruitment of macrophages to the islets. Soluble mediators produced by activated macrophages play a critical role in the destruction of residual beta cells. A single amino acid at position 776 of the EMC viral genome controls the diabetogenicity of the virus. In contrast, KRV causes autoimmune type 1 diabetes in diabetes-resistant BioBreeding (DR-BB) rats without direct infection of beta cells. Macrophages play an important role in the development of diabetes in KRV-infected DR-BB rats. As well, KRV infection preferentially activates effector T cells, such as Th1-like CD45RC(+)CD4(+) T cells and CD8(+) T cells, and downregulates regulatory T cells, such as Th2-like CD45RC(-)CD4(+) T cells. This results in the breakdown of the immune balance, contributing to the development of diabetes in KRV-infected DR-BB rats.